Immature thymocytes isolated using a sialic acidspecific lectin are unresponsive to concanavalin A

Abstract

Mouse thymocytes, which are approximately 90% immature cortical cells, are low in surface sialic acid when compared with more mature cortisone-resistant, presumably medullary, thymocytes and peripheral T lymphocytes. Thus, medullary thymocytes bind and are agglutinated by the N-acetylneuraminic acid-specific lectin, lobster agglutinin 1 (LAgl), whereas cortical thymocytes are not agglutinated by this lectin. It is demonstrated herein that mouse cortical thymocytes, purified using LAgl, do not respond to the mitogenic effects of concanavalin A (Con A). The lack of response to this lectin is not due to depletion of macrophages, since addition of macrophages does not restore the response. Populations of LAgl-negative thymocytes can be made to respond weakly to Con A by the addition of interleukin 2, but this response appears to be due to the presence of a few contaminating LAg1-binding thymocytes since it is abolished by treatment of the cells with rabbit anti-LAgl serum plus complement. Therefore highly purified cortical thymocytes not only cannot respond to Con A, but also they cannot be induced to respond by the addition of the T cell growth factor, interleukin 2. Thymocytes isolated by a single criterion, that is by virtue of their low amount of surface sialic acid, appear to be a truly immature population of thymocytes.

Low sialic acid‐bearing mouse thymocytes do not express helper T cell properties in vitro

“…The major population of mouse thymocytes bears low levels of surface sialic acid [l], and thus binds the galactose-specific lectin, peanut agglutinin (PNA), but not the sialic acid-specific lectin, lobster agglutinin 1 (LAgl) [2-61. In a previous report, we described the use of LAgl to isolate low sialic acid-bearing, presumably cortical, thymocytes; these LAgl-thymocytes do not demonstrate a significant proliferative response to the T cell mitogen concanavalin A (Con A), even when cultured in the presence of interleukin 2 (IL2) [6]. These results are in agreement with recent reports by others in which carefully isolated populations of PNAf cortical thymocytes were used [ 7 , 81. In this report we further characterize LAgl-thymocytes as cells which do not produce or respond to IL2 when stimulated with Con A.…”

“…Thymocytes were separated into LAgl' and LAgl-subpopulations by a modification of the procedure described earlier [6]. Thymocyte suspensions of 2 x 108-20 x 10' cells in BSS, lo9 cells/ml, were incubated with 0.2-2 ml LAgl (0.1 ml LAgl at a protein concentration of 2-5 mgiml per 10' cells).…”

Thymic cytotoxic T lymphocytes are primed in vivo to minor histocompatibility antigens.

“…However, this finding has little bearing on the sensitivity of responding cells at the time of antigen injection, for the necessary 10 d interval between priming and culture is more than enough time for cortisone-sensitive cells to become cortisone resistant (31). It appears that even in the presence of an excess of helper factors, cells of minority ("medullary") phenotype comprise the vast majority of thymocytes that proliferate or differentiate to CTL in the presence of Con A (23,32) or allogeneic stimulator cells (24). It must be stressed that this minority phenotype has been defined by parameters other than intrathymic location.…”

Interleukin 4 is a growth factor for activated thymocytes: possible role in T-cell ontogeny.

“…The erythrocytes were lysed by hypotonic shock. In some experiments, thymocytes were fractionated on the basis of their binding to PNA (Sigma) as described (16). Briefly, cells were agglutinated with PNA (20 Ag/ml) in balanced salt solution for 30 min at 37TC.…”