Kinetic analysis of thymocyte attachment to thymus stromal cells in culture by using phase‐contrast and scanning electron microscopy

Abstract

Direct cellular contact between thymocytes and thymus stromal cells within the thymus appears to contribute to the maturation of thymocytes. Thymocyte-stromal cell complexes, formed in vivo, have been isolated by others and postulated to play a role in T-cell differentiation. These previous studies have been hampered, however, by a time-consuming isolation procedure from which only small numbers of these complexes are recovered. We have examined a model to study thymocyte-stromal cell complexes in vitro in which thymocytes are added to primary cultures of thymus stromal cells. In the present study, we found that thymocytes were histotypically selective in their attachment to thymus stromal cells. We also investigated the kinetics of thymocyte attachment to these thymus stromal cells. Cultures were examined at selected time intervals from 5 min through 3 days of incubation. Thymocyte attachment to stromal cells was a biphasic interaction, with maximum surface attachment at 15 min of cocultivation, followed by migration of thymocytes into the cultures. Morphological studies were confirmed by using 3H-leucine-labeled thymocytes and liquid scintigraphy. With increased time in culture, thymocytes became amoeboid and migrated between the layers of stromal cells where thymocyte mitotic figures were seen at 4 and 8 hr. In some cases it appeared that stromal cells, which often grew two to three cell layers deep, played an active role in enclosing thymocytes within the cultures. Large numbers of viable thymocytes were observed in the cultures at 24 hr. The number of thymocytes then decreased progressively on days 2 and 3, when relatively few were found within the layers of the culture.

Morphological analysis of the cellular interaction between thymocytes and a thymic stromal cell line (TEL‐2)

“…The ultrastructure of the binding site indicated closely apposed membranes of both cells (data not shown). This finding may suggest the presence of specific receptors for cellular recognition and binding between thymocytes and TEL-2 cells (LaRochelle and Jones, 1989). A noticeable property of the cultures of TEL-2 cells was that they formed numerous "round spaces."…”

“…Rapid intercellular adhesion seems to be common in the cellular interaction of thymocytes and thymic stro-ma1 elements (LaRochelle and Jones, 1989). Many thymocytes became attached to the surface of the TEL-2 cells shortly after cocultivation.…”

Morphological study of thymus stromal cells (TEL-2 cell) which play a role in the elimination of double positive immature thymocytes by phagocytosis

“…In a follow-up study with SEM and transmission electron microscopy (TEM), Hirata et al (1991) have shown that as an initial event for the process of the selective elimination of DP thymocytes from the culture, the attachment between the thymocyte and TEL-2 cell leads to a migration of thymocytes between the TEL-2 processes. Similar cellular interactions between thymocytes and thymic stromal elements have been described in vitro (LaRochelle and Jones, 1989). It is, however, still open to question concerning the cytological events which occur as a sequelae of the initial attachment.…”

“…7d), followed the ingestion of thymocytes by TEL-2 cells. The ultrastructure of these binding sites may suggest the presence of specific receptors for cellular recognition between thymocytes and thymus stromal cells (TEL-2 cells) (LaRochelle and Jones, 1989;Hirata et al, 1991).…”