SummaryTo seek direct evidence for the notion that stem cells in the thymus need to be constantly replenished from the bone marrow (BM), fetal (day 15) thymuses from normal BALB/c mice were grafted into T and B cell-deficient C.B-17 SCID mice (both H-2 d, I-E+). The thymus grafts in these mice showed normal thymopoiesis for the first 3 wk postgrafting but then developed sudden atrophy with near complete loss of CD4+8 + cells by 4-5 wk. Such atrophy was not seen when the thymus-grafted mice were cotransplanted with normal BM cells. The lymph nodes of SCID mice receiving thymus grafts alone contained mature T cells but virtually no B cells. This lack of B cells was associated with aberrant I-E-restricted Vfl deletion: the depletion of Vfl3 + and Vfl5 + T cells was near complete, whereas Vflll + cells showed only marginal depletion.N 'eonatal thymus grafts (TG) 1 placed in normal or thymectomized mice undergo considerable enlargement and remain functional for prolonged periods (1, 2). The cells proliferating in the TG are initially of donor origin but are rapidly replaced by host-derived cells. This transition is complete by 1 mo posttransfer. Similarly, intrathymic injection of CD4-8-thymic stem cells results in only transient thymopoiesis by donor-derived cells (3-5). These data imply that the self-renewing capacity of thymic stem cells is limited and that maintenance of thymic function depends on a continuous inflow of stem cells from the bone marrow (BM).A corollary to this notion is that depriving the thymus of an exogenous source of functional stem cells would lead to rapid atrophy. This prediction can be tested by placing TG in SCID mice (6). These mice have an autosomal recessive mutation that prevents the formation of T and B cells; T cell differentiation in SCID mice proceeds only to the stage of CD4-8-thymocytes. We show here that day 15 BALB/c fetal TG placed in H-2-compatible adult C.B-17 SCID mice exhibit normal thymopoiesis for 3 wk but then undergo sudden atrophy with selective disappearance of CD4 + 8 + cells. The LN of these mice contain large numbers of T cells but virtually no B cells. The absence of B cells is associated with aberrant Vfl deletion.1 Abbreviations used in this paper: B/c, BALB/c; TG, thymus grafts or thymus grafted.
Materials and MethodsM/ca C.B-17 SCID mice (6) and other mice were obtained from the breeding colony of the Scripps Research Institute. SCID mice were maintained under pathogen-free conditions. Note that C.B-17 and BALB/c are identical except for an Ig allotype difference.Thymus Grafting. Day 15 fetal thymuses from timed-pregnant mice were inserted under the kidney capsule (four lobes/recipient) of young adult SCID mice using standard procedures. Some of the grafted mice received ",,5 x l& T cell-depleted normal donor BM cells (7) intravenously within 1 d of grafting.Flow Cytometry. Thymus and/or LN cells were stained for expression ofCD4 (GK1.5), 3.168, or YTS 169), c~/fl TCR (H57-597), IL-2R (7D4), B220 (14.8), CD45RB (23G2), Thy-1.1 (19E12), and TCR V33 (KJ25), -...
The work presented here attempts to consolidate our knowledge on cellular transcriptome and proteome. It takes into account that a typical activated cell (lymphocyte) contains 40 000 mRNA molecules at any time, and it represents about 5000 different molecular species of transcripts. Such a cell has about 1 000 000 000 protein molecules, some of them being present at 10 000 000 copies while others at a very low copy number (say 1 to 10 copies per cell). By studying cell free expression of individual cDNA clones (or pools of known complexity) we address to those rare molecular components that will remain undetected by the current analytical means. For our analysis we use cell free translation systems (wheat germ or rabbit reticulocyte origin) and we study polypeptide products originating from intact, or restriction endonuclease-treated cDNA clones. We conclude that in most instances expressed genes yield transcript(s) that translate into several, and often very numerous families of polypeptide species. In our ISODALT two-dimensional gel system we characterize the proteomic profile of the clonal polypeptide families in terms of their molecular mass, charge, multiple products, and appearance.
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