Seven representative T cell tracks (colored lines) are shown for both and a time stamp (hr:min:sec:msec) is placed in the upper right corner of each frame

Seven representative T cell tracks (colored lines) are shown for both and a time stamp (hr:min:sec:msec) is placed in the upper right corner of each frame. CD5 is a negative regulator of TCR-mediated signaling (25C28), we anticipated that this change would result in hyperresponsiveness to antigenic stimulation. In contrast, we found that the ability of OT-II cells to proliferate in response to Act-mOVA-expressing DCs injected on the same day as the OT-II cells was JZL195 lower in MHC class II-deficient compared with MHC class II-sufficient recipients. This defect became more pronounced when the OT-II cells were deprived of MHC class II contact for 24 h, and was most profound with additional days of deprivation (Fig. 1= 0= number of cell divisions. The graph shows data from spleens of CD4-deficient (filled squares) and MHC class II-deficient (open circles) recipients. Each data point shows the mean (SD) of four animals. These data are representative of five independent experiments. ( 0.001) or 0 days of deprivation (46%, 0.01). In contrast DCs not contacting OT-II cells in the MHC class II-sufficient environment (Fig. 2 0.001, for each) but not with 0 days of MHC class II deprivation. Thus, when OT-II cells are exposed to an Klf2 MHC class II-deprived environment, the lack of contact of the cells with DCs correlates with decreased CD69 expression (Fig. 1 0.05). Although the functional relevance of large DC/T cell clusters has not been clearly demonstrated, this has been shown to correlate with CD4 T cell effector function in similar systems (i.e., delayed-type hypersensitivity response) (23, 30, 31). These visualization studies strongly suggest that self-ligand-deprived T cells have an impaired ability to contact and/or adhere to antigen-bearing DCs. Open in a separate window Fig. 2. OT-II T cells fail to form clusters with antigen-bearing DCs after transfer into MHC class II-deficient recipients. CFSE-labeled OT-II cells were adoptively transferred into MHC class II-deficient or CD4?/? mice on the day of (day 0) (and and and 0.001. MHC Class II Deprivation Leads to Reduced CD4 T Cell Motility in Secondary Lymphoid Tissue. We JZL195 next wanted to determine why, despite the presence of antigen-bearing DCs within the paracortex of the lymph node, the CD4 T cells fail to cluster with the DCs and proliferate. We hypothesized that there was a defect in motility of the CD4 T cells. A defect in motility could result in impaired DC contact and reduced TCR signaling JZL195 and proliferation. We performed video microscopy on explanted lymph nodes to determine OT-II cell velocity and path of movement after transfer into MHC class II-deficient and wild-type animals. We observed a profound defect in the motility of OT-II cells deprived of MHC class II molecules in lymph nodes in the absence of antigen (SI Movie 1) compared with OT-II cells transferred into an MHC class II-sufficient environment (SI Movie 2). In fact, the longer the cells resided in the MHC class II-deficient animals, the more profound the defect, in both their velocity and their tracking path (Fig. 3). As an additional measure of the sessile nature of the cells, we analyzed the numbers of OT-II cells in the blood and the spleen 7 days after transfer. Compared with MHC class II-sufficient recipients, MHC class II-deficient recipients had fewer OT-II cells in the blood despite similar numbers in the spleens (SI Fig. 10). These data suggest that motility, before antigen encounter, is an essential component of CD4 T cell activation and that it is influenced by contact with MHC class II molecules. Open in a separate window Fig. 3. MHC class II deprivation impairs CD4 T cell motility. (and projections of the imaging volume (200 225 75 m). Seven representative T cell tracks (colored lines) are shown for both and a time stamp (hr:min:sec:msec) is placed in the upper right corner of each frame. (and (19). Indeed, our studies support the notion that continuous stimulation from self ligands expressed by APCs is likely to be of considerable significance for the functional responsiveness of T cells toward cognate antigens (Fig. 1and SI Fig. 7). However, with more prolonged deprivation (i.e., days rather than hours) we found a defect in the expression of CD69 (Fig. 1challenge of antigen, the previously MHC class II-deprived OT-II cells proliferated as well as the control OT-II cells that were not previously MHC class II-deprived. These data suggest that the T cells were unable to effectively contact antigen-bearing DCs that is not evident in the assays JZL195 because the T cells are able to contact APCs passively. However, the defective motility which we report for CD4 T cells deprived of MHC class II molecules may result from loss of either a specific interaction (involving TCR recognition) and/or a general interaction (for example, CD4 binding). Our.


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