T Regulatory Cell Kinetics Are Altered in a Target Organ of Chronic Gvhd, Resulting in a Low T Regulatory to T Effector Memory Cell Ratio

T regulatory cells kinetics are altered in a target organ of cGVHD, resulting in a low T regulatory to T effector memory cell ratio

Chronic graft-versus-host disease (cGVHD) is a major cause of morbidity and mortality following allogeneic hematopoietic stem cell transplant (AHSCT).  Although current therapies largely target T cell proliferation, in vivo T cell kinetics of division, death and trafficking have been difficult to evaluate.  We employed deuterated water labeling with subsequent GC-MS/MS enrichment detection to measure in vivo T cell kinetics after AHSCT in a mouse model of cGVHD. We investigated CD4 T cell sub-population kinetics of cell division, death and trafficking, in non-target and target tissues of cGVHD.   Early in the disease course, spleens of AHSCT recipients showed a high number of CD4+ T regulatory (Treg) and T effector memory (Tem) cells.  In vivo cell division kinetics of these two cell types were high and similar to each other, and slowed as cGVHD progressed.   Meanwhile, Tem cell loss from the spleen was significantly faster than that of Treg cells, likely as a result of increased trafficking.  In support of trafficking was the finding of elevated Tem numbers in peripheral blood of AHSCT recipients despite a relative lymphopenia early in the disease with subsequent disappearance from the circulation once cGVHD was clinically evident.  Furthermore, overall Tem cell numbers in the host were significantly higher than the number present in the graft, but distributed to target rather than non-target tissues.  Target sites of cGVHD (skin, gut and liver) had several fold greater number of Tem cells compared to controls both early and late in disease.  The number of Tem cells in target sites was maintained as cGVHD progressed, and the Treg to Tem ratio in target tissues was significantly lower than that of the controls at all times.  Cell division of Tem and Treg cells was high in the liver, and remained high for the Treg cells as disease progressed.  Meanwhile, Treg cells underwent faster loss from the liver, likely due to cell death given that the total number of Treg cells did not change.  Our findings shed light on in vivo behavior of Treg and Tem cells, T cell subtypes critical to the biology of cGVHD.  Furthermore, our methodology will be translated to investigate in vivo T cell kinetics in patients undergoing AHSCT as deuterated water labeling is safe, not toxic and non-radioactive.

Figure 1.  Total number of CD4+ Treg and Tem cells in the syngeneic vs. allogeneic recipients.  Shown are total number of CD4+ Treg and Tem cells in the graft (day 0), and at subsequent time points following AHSCT for syngeneic (syn) recipients, identical host recipient pairs; and allogeneic (allo) recipients, with minor antigen mismatch between host and donor with resultant cGVHD induction.  Day+14 is a time point early post AHSCT, while day +28 represents well-established cGVHD.