Visualizing thymocyte motility using 2-photon microscopy

Department of Molecular and Cell Biology, University of California, Berkeley, CA 94720, USA.
Immunological Reviews (Impact Factor: 10.12). 11/2003; 195(1):51-7. DOI: 10.1034/j.1600-065X.2003.00069.x
Source: PubMed


Our view of a thymocyte based on its behavior in tissue culture and appearance in fixed tissue sections was of a round sessile cell. Its travel through the thymus might occur slowly, perhaps even passively, leaving it in contact with the support cells that happened to be in its immediate environment. However, when we got our first look at the behavior of thymocytes in a 3D cellular stromal cell environment, that picture changed dramatically. Instead we found that thymocytes are actively crawling, allowing them to explore their environment over relatively long distances and interact with peptide-major histocompatibility complex (pMHC)-bearing thymic stromal cells in both dynamic and stable modes. In this review, we discuss the implications of thymocyte motility for T-cell repertoire selection and for the mechanisms that determine the spatial organization of thymocyte subsets within the thymus.

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    • "Immunoimaging techniques have now been applied to isolated organ and tissue preparations, including lymph node [6] [7] [8] [9] [10] [11] [12] [13], thymus [14] [15] [16] [17], spleen [18], intestinal tissue [19], skin [20], and in brain and spinal cord slices [21] [22]. In addition, for the first time, immune cells have been imaged in living, anesthetized mice, using intravital preparations of lymph nodes [5] [23] [24]. "
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    ABSTRACT: Cellular interactions in lymphoid organs initiate the immune response and determine its outcome. Using two-photon microscopy in the lymph node, several groups have begun to investigate the motility characteristics and interactions among T lymphocytes, B lymphocytes, and dendritic cells (DC) in lymphoid organs. In the first "close encounter", T cells of a particular antigen specificity interact with antigen-bearing dendritic cells and begin to activate. Activation of both CD4+ and CD8+ T cells evolves through several stages; from transient interactions to stable clusters and later to dissociation and proliferation of T cells (clonal expansion). The second "close encounter" requires that antigen-engaged B cells become accessible to T cells by directed migration to the edge of the follicle. T cells and B cells then pair up and waltz together for an extended period, while helper T cells provide signals for B cells to differentiate into plasma cells. In this topical review, we compare the activation choreography of CD4+ T cells interacting first with dendritic cells, and then with B cells, during initiation of the humoral immune response.
    Seminars in Immunology 01/2006; 17(6):442-51. DOI:10.1016/j.smim.2005.09.001 · 5.17 Impact Factor
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    • "However, the scanning time of DCs not presenting cognate antigen seems more likely to lie at the lower end of the range. This is supported by the observation that the peak T cell migration rate in ex vivo lymph nodes and foetal thymic organ culture observed using two-photon microscopy was in excess of 25 m min −1 , which would suggest a scanning rate of at least 1 APC/min [33] [42]. It is therefore possible that n 1 may exceed 20,000, assuming a 2-week negative selection window and 1 min/APC encounter. "
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    ABSTRACT: The activation of a T cell is a stochastic process, and depends on the integrated strength of signals 1 and 2 resulting from its encounter with an antigen presenting cell. The net outcome of thymic selection and peripheral circulation over many such encounters in the presence of mechanisms for both central and peripheral tolerance is difficult to deduce by intuition alone. We therefore introduce a simple mathematical model that allows us to explore the roles and interaction of different thresholds, costimulation and anergy, as well as make predictions about expected immune system behaviour. We show that stochastic activation in the context of repeated encounters results in lowering the apparent activation threshold for T cells. This effect may contribute significantly to the efficiency of negative selection, although a low avidity subset of auto-reactive thymocytes can still be exported. A simple peripheral mechanism for peripheral tolerance is shown to be highly effective at dealing with this low avidity subset. Finally, the trade-offs between sensitivity and specificity are examined.
    Journal of Computational and Applied Mathematics 12/2005; 184(1-184):101-120. DOI:10.1016/ · 1.27 Impact Factor
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    • "We must assume in this case that transitional cells enter the thymic medulla, and then “zoom” around the medulla at high speed. Recent studies using multiphoton microscopy of T cells in lymph node and thymic organ cultures have shown that they travel at an average velocity of 14 μm per min, and can reach peak velocities of 40 μm per min (33–36). If newly generated medullary T cells also locomote like this they could potentially interact with 1,000–5,000 Mtecs in 1 d. "
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    ABSTRACT: Intrathymic expression of tissue-specific antigens (TSAs) by medullary thymic epithelial cells (Mtecs) leads to deletion of autoreactive T cells. However, because Mtecs are known to be poor antigen-presenting cells (APCs) for tolerance to ubiquitous antigens, and very few Mtecs express a given TSA, it was unclear if central tolerance to TSA was induced directly by Mtec antigen presentation or indirectly by thymic bone marrow (BM)-derived cells via cross-presentation. We show that professional BM-derived APCs acquire TSAs from Mtecs and delete autoreactive CD8 and CD4 T cells. Although direct antigen presentation by Mtecs did not delete the CD4 T cell population tested in this study, Mtec presentation efficiently deleted both monoclonal and polyclonal populations of CD8 T cells. For developing CD8 T cells, deletion by BM-derived APC and by Mtec presentation occurred abruptly at the transitional, CD4high CD8low TCRintermediate stage, presumably as the cells transit from the cortex to the medulla. These studies reveal a cooperative relationship between Mtecs and BM-derived cells in thymic elimination of autoreactive T cells. Although Mtecs synthesize TSAs and delete a subset of autoreactive T cells, BM-derived cells extend the range of clonal deletion by cross-presenting antigen captured from Mtecs.
    Journal of Experimental Medicine 11/2004; 200(8):1039-49. DOI:10.1084/jem.20041457 · 12.52 Impact Factor
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