Kkm. Verbanac et al., A ROLE FOR TRANSFORMING GROWTH-FACTOR-BETA IN THE VETO MECHANISM IN TRANSPLANT TOLERANCE, Transplantation, 57(6), 1994, pp. 893-900
We have studied the veto cell-mediated induction of transplant toleran
ce by allogeneic donor bone marrow cells and have achieved kidney allo
graft tolerance in a preclinical rhesus monkey model. Here we extend t
hese studies to investigate the veto mechanism of CTLp suppression and
the role of CD8 and TGF-beta in these events. Infusion of DR-/dim don
or BMC into RATG-treated rhesus monkeys induced functional deletion of
donor-specific CTLp and prolongation of kidney allograft survival, wh
ereas depletion of the CD8+ subset from BMC ablated these effects. A r
ole of CD8 in the veto effect was further implicated by rhesus MLR-ind
uced CML experiments in which pretreatment of normal responder cells w
ith MAb to MHC class I, the natural ligand of CD8, blocked the suppres
sive activity of allogeneic BMC. In addition, pretreatment of the BMC
with anti-CDS MAbs blocked strong veto activity significantly, suggest
ing that CD8 functions as an accessory or adhesion ligand. In contrast
, anti-CD8 treatment significantly enhanced weak BMC-mediated veto act
ivity, suggesting that CD8 might additionally serve as a signal transd
ucer to increase veto activity, perhaps by the induction of cytokine r
elease. The cytokine TGF-beta was studied because it has immunosuppres
sive properties that are shared by veto cells. Human TGF-beta, like BM
C veto cells, inhibited MLR-induced CML in a dose-dependent manner, an
d anti-TGF-beta Ig relieved the BMC-mediated veto suppressive effect.
Active TGF-beta was detected only in the supernatants of CML cultures
containing BMC. Pretreatment of BMC with L-leucyl-leucine methyl ester
(Leu-leu-OMe), which eliminates cytotoxic precursor and effector lymp
hocytes and monocytes, did not affect levels of active TGF-beta. In pr
evious studies, the veto effect of BMC was also shown to be Leu-leu-OM
e-resistent. Finally, treatment of isolated DR-/dim BMC cultures with
anti-CD8 elicited TGF-beta secretion, whereas anti-CD2 or anti-CD3 had
no effect. When isolated after stimulation with anti-CD8, only the CD
8+ subset of DR-dim BMC produced detectable levels of active TGF-beta.
In summary, these studies demonstrate that CD8 functions as an immuno
regulatory molecule in veto effects by freshly isolated rhesus BMC and
suggest that CD8-ligand interactions may induce low-level secretion o
f TGF-beta to mediate or facilitate the veto mechanism of CTLp inactiv
ation in a paracrine manner.