Cross-Linking the TCR Complex Induces Apoptosis in CD4+8+Thymocytes in the Presence of Cyclosporin A

Although it is generally agreed that TCR ligation is a minimal requirement for negative selection in the CD+8+double-positive (DP) thymocyte subset, the costimulatory requirements and specific signaling events necessary to induce apoptosis are not well defined. We have explored the consequences of cross-linking CD3/TCR complexes on thymocytes from H-Y TCR transgenic (Tg) mice. In agreement with previous reports, we demonstrate that culturing DP thymocytes with plate-bound anti-TCR antibody induces downregulation of CD4 and CD8 and upregulation of CD69 expression. Nevertheless, the activated cells did not undergo apoptosis, as determined by viable cell recoveries and by quantitation of DNA fragmentation using the TUNEL assay. However, specific depletion of the DP subset occurred within 24 hr when thymocytes were incubated in the presence of both anti-TCR and the immunosuppressant cyclosporin A (CsA). CsA also induced depletion of anti-CD3 stimulated normal DP thymocytes. Using mice homozygous for thelprorgldmutation, we also have shown that Fas/Fas ligand interactions are not involved in the CsA-induced death of TCR-stimulated DP thymocytes. These data verify that TCR cross-linking alone is insufficient to induce apoptosis of DP thymocytes and further suggest that TCR stimulation activates a CsA-sensitive protective pathway that interferes with signaling events leading to apoptosis in DP thymocytes.

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In vivo and in vitro clonal deletion of double-positive thymocytes.

Journal of Experimental Medicine ◽

10.1084/jem.175.5.1307 ◽

1992 ◽

Vol 175 (5) ◽

pp. 1307-1316 ◽

Cited By ~ 114

Author(s):

N J Vasquez ◽

J Kaye ◽

S M Hedrick

Keyword(s):

T Cells ◽

T Cell ◽

Cytochrome C ◽

Cyclosporin A ◽

Negative Selection ◽

Double Positive ◽

Antigen Dose ◽

Mhc Molecules

To study the processes of thymic development, we have established transgenic mice expressing and alpha/beta T cell antigen receptor (TCR) specific for cytochrome c associated with class II major histocompatibility complex (MHC) molecules. The transgenic TCR chains are expressed by most of the thymocytes in these mice, and these cells have been shown to efficiently mature in association with Ek- and Ab-encoded class II MHC molecules. This report describes a characterization of the negative selection of these transgenic thymocytes in vivo that is associated with the expression of As molecules. Negative selection by As molecules appears to result in the deletion of a late stage of CD4/CD8 double-positive thymocytes in that there is a virtual absence of transgenic TCR bearing CD4 single-positive thymocytes. This phenotype is accompanied by the appearance of CD4/CD8 double-negative thymocytes and peripheral T cells that are functionally antigen reactive. The process of negative selection has also been investigated using an in vitro culture system. Upon presentation of cytochrome c by Eb-expressing nonthymic antigen-presenting cells, there occurs an antigen dose-dependent deletion of the majority of CD4/CD8 double-positive thymocytes. In contrast, presentation of Staphylococcal enterotoxin A by Eb in vitro results in minimal deletion of double-positive thymocytes. In addition, we use this in vitro model to examine the effects of cyclosporin A on negative selection. In contrast to its effects on mature T cells, and the findings of others in vivo, cyclosporin A does not inhibit antigen-induced deletion of double-positive thymocytes. Finally, a comparison of the antigen dose responses for thymocyte deletion and for peripheral T cell activation indicates that double-positive thymocyte recognition is more sensitive than mature T cells to antigen recognition.

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Activated CD4+ T cells enriched from spleen or colon induce apoptosis in intestinal epithellal cells via fas/fas ligand interactions

Gastroenterology ◽

10.1016/s0016-5085(01)80949-5 ◽

2001 ◽

Vol 120 (5) ◽

pp. A192-A192

Author(s):

H TAKAISHI ◽

T DENNING ◽

K ITO ◽

R MIFFLIN ◽

P ERNST

Keyword(s):

T Cells ◽

Cd4 T Cells ◽

Fas Ligand ◽

Ligand Interactions

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CD45-null transgenic mice reveal a positive regulatory role for CD45 in early thymocyte development, in the selection of CD4+CD8+ thymocytes, and B cell maturation.

Journal of Experimental Medicine ◽

10.1084/jem.183.4.1707 ◽

1996 ◽

Vol 183 (4) ◽

pp. 1707-1718 ◽

Cited By ~ 269

Author(s):

K F Byth ◽

L A Conroy ◽

S Howlett ◽

A J Smith ◽

J May ◽

...

Keyword(s):

Signal Transduction ◽

T Cell ◽

B Cells ◽

Transgenic Mice ◽

B Cell ◽

T Cell Development ◽

Cell Development ◽

Cross Linking ◽

Thymocyte Development ◽

Double Positive

The CD45 transmembrane glycoprotein has been shown to be a protein phosphotyrosine phosphatase and to be important in signal transduction in T and B lymphocytes. We have employed gene targeting to create a strain of transgenic mice that completely lacks expression of all isoforms of CD45. The spleens from CD45-null mice contain approximately twice the number of B cells and one fifth the number of T cells found in normal controls. The increase in B cell numbers is due to the specific expansion of two B cell subpopulations that express high levels of immunoglobulin (IgM) staining. T cell development is significantly inhibited in CD45-null animals at two distinct stages. The efficiency of the development of CD4-CD8- thymocytes into CD4+ CD8+ thymocytes is reduced by twofold, subsequently the frequency of successful maturation of the double positive population into mature, single positive thymocytes is reduced by a further four- to fivefold. In addition, we demonstrate that CD45-null thymocytes are severely impaired in their apoptotic response to cross-linking signals via T cell receptor (TCR) in fetal thymic organ culture. In contrast, apoptosis can be induced normally in CD45-null thymocytes by non-TCR-mediated signals. Since both positive and negative selection require signals through the TCR complex, these findings suggest that CD45 is an important regulator of signal transduction via the TCR complex at multiple stages of T cell development. CD45 is absolutely required for the transmission of mitogenic signals via IgM and IgD. By contrast, CD45-null B cells proliferate as well as wild-type cells to CD40-mediated signals. The proliferation of B cells in response to CD38 cross-linking is significantly reduced but not abolished by the CD45-null mutation. We conclude that CD45 is not required at any stage during the generation of mature peripheral B cells, however its loss reveals a previously unrecognized role for CD45 in the regulation of certain subpopulations of B cells.

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