INFLUENCE OF CATHPESIN-L ON CD4+ REGULATORY T CELLS PERIPHERAL

CAMICIA GABRIELA; N BADANO; MAGLIOCO ANDREA; COSTA HÉCTOR; I PIAZZON; I NEPOMNASCHY

Congreso; Congreso Franco Argentino de Inmunología; 2010

CD4+CD25+Foxp3+ regulatory T cells play a pivotal role in the maintenance of peripheral tolerance and immune homeostasis. We had previously shown that cathepsin-L deficient mice (CTSLnkt) show increased absolute number of CD4+ regulatory T cells in peripheral lymph nodes (LN) whereas the number of CD4+Foxp3+ thymic cells was shown to be decreased. In this study, we investigated the impact of cathepsin-L on CD4+ regulatory T cell peripheral homeostasis by analyzing CD4+CD25+ (Treg) cell turnover in CTSLnkt mice. To determine the proliferation rate of LN Treg subsets, 5-bromo-2- deoxyuridine (BrdU) was injected for 7 days. FACS analysis showed that both the CTSLnkt Treg and CD4+CD25- subsets showed a significant increase in the % of BrdU+ cells as compared to wild type (mean % BrdU+/Treg cells ± DS: 22±3 vs 12±1, p<0.005, n=4. Mean % BrdU+/CD4+CD25- cells ± DS: 10±1 vs 4.0±0.1, p<0.001, n=4). To evaluate the % of Treg cells undergoing apoptosis annexin V staining was used. An increase both in the % of apoptotic Treg and CD4+CD25- cells was observed in the LN of mutant mice (mean % annexin V+/Treg cells ± DS: 53±3 vs 43±3, p<0.005, n=4. Mean % annexin V+/CD4+CD25- cells ± DS: 35±2 vs 25±2, p<0.001, n=4). Notably, both mutant and wild type Treg subsets showed increased apoptosis as compared to their CD4+CD25- counterparts. Our results show that the CTSLnkt mutation causes increases in the proliferation but also in the apoptosis levels of both Treg and CD4+CD25- cells. The fact that mutant Treg and CD4+CD25- cells showed increases in proliferation but also in the apoptosis levels, does not support that differences in the proliferation vs apoptosis balance of Treg may be involved in the increases of peripheral Treg numbers in CTSLnkt mice, thus raising the possibility that conversion of CD4+CD25- into CD4+CD25+ cells could be involved. In support of this hypothesis, increased levels of TGF-b were found in cultures of LN CTSLnkt cells using both RT-PCR and ELISA assays.