Assess Cytotoxic, Helper and Regulatory T Cells (Treg) populations, before and after AHSCT in patients with T1D and to correlate with clinical outcomes.

Peripheral Blood Mononuclear Cells collected from 11 T1D patients were analyzed by Flow Cytometry to identify T Cytotoxic (CD3+CD8+), T Helper (CD3+CD4+) and Treg populations (CD3+CD4+CD25+FOXP3+). Results were retrospectively correlated with the patients'clinical outcomes of short (under 3-years) and long insulin independence (over 3-years).

Most patients were male (54.5%) with a median age of 18.6-years (16‒23). After transplantation, patients became independent of exogenous insulin and had an increase in peptide C levels for up to a year after AHSCT. Analyzing all patients, TCD8+ cells increased when comparing baseline, before AHSCT, to 60, 100, 270 and 360 days post treatment (p < 0.01). TCD4+ frequency was decreased at 60 (p < 0.05) and 100 (p < 0.001) days after AHSCT, when comparing baseline. The frequency of Tregs (CD4+ CD25+ Foxp3+) increased at 60-days post-AHSCT compared to baseline (p < 0.05) and decreased at 180-days from 60-days post treatment (p < 0.05). When analyzing insulin independence groups separately, it was observed that short-term independence patients had higher TCD8+ frequency in timepoints less than 6-months post transplant (p < 0.05), and lower TCD4+ frequency in the same time point (p < 0.05), compared to long-term patients. Also, long-term insulin independence patients had higher baseline Treg frequency (p < 0.01) and these levels maintained higher in periods up to 6-months (p < 0.05), compared to the short-term group.

When analyzing all patients, while there is an increase in T Cytotoxic lymphocyte populations in periods up to 6-months after transplantation, T Helper cells decrease at the same time points. Thus, our results demonstrate the dynamics of TCD4 and TCD8 cell reconstitution during the homeostatic proliferation period following profound lymphopenia caused by the conditioning regimen. Additionally, there is an increase in Treg frequency at early time points after the transplant, which may contribute to immune tolerance during the period of proliferation of residual or introduced cells in the graft. During studies based on the duration of insulin independence, we observed that long-term insulin-independent patients have a higher Treg frequency at baseline compared to short-term insulin-independent patients, suggesting that these patients might have a more tolerant initial immune system. The long-term group maintains higher Treg percentage in periods up to six months, which could contribute to the favorable clinical response. Also, long-term insulin-independent patients show less cytotoxic cells and more helper cells in periods less than 6-months, which may have contributed to the clinical outcome, since T1D is mainly associated with a cytotoxic auto-reactive response.

The results of this study corroborate with previous findings in regards to the immune system reconstitution after AHSCT. The better responding group have, in some time points, lower frequencies of TCD8+ cells, that mediate auto-reactive inflammation, and higher frequencies of Treg,known for promoting self-tolerance. This suggests that the clinical success of AHSCT for T1D, and potentially other autoimmune disorders, is related to the profile of the patients’ reestablished immunity, emphasizing the importance of immunological monitoring studies.