Loss of adenosine deaminase activity inhibits beta selection in murine fetal thymic organ culture

Murine fetal thymic organ cultures (FTOC) treated with the ADA inhibitor 2′-deoxycoformycin (dCF) were used as a model system to investigate the mechanism by which T cell differentiation is inhibited in adenosine deaminase (ADA)-deficient humans. The metabolic abnormalities caused by dCF were similar to those in ADA-deficient patients and ADA knock-out mice. They included inhibition of S-adenosyl homocysteine hydrolase and elevation of intrathymic adenosine, deoxyadenosine, and total deoxyadenylates. In FTOC established on gestational day 14 with 5 μM dCF and harvested 6 days later, thymocyte differentiation was blocked in the CD4-CD8- compartment at the CD44-CD25+ to CD44-CD25- transition. However, neither TCR Vβ chain gene rearrangement nor pre-TCRα expression were inhibited. Furthermore, the effects of dCF treatment were not overcome by rearranged TCR α and β transgenes. These results are consistent with an inhibition of lck-mediated signaling through the pre-TCR; i.e., β selection. To test this hypothesis, we analyzed the effects of dCF upon FTOC with Rag-2-/- thymuses in the presence of anti-CD3. As expected, under control conditions Rag-2-/- thymocytes differentiated to the double positive stage with anti-CD3 treatment. However, in the presence of dCF, progression past the double negative stage was inhibited. These data indicate that the metabolic derangements which occur when ADA is absent interfere with pre-TCR signaling in the developing thymus.

Duke Scholars

Author Michael Steven Hershfield Medicine, Rheumatology and Immunology