Influence of cathepsin-L on CD4+ regulatory T cells peripheral homeostasis.”

CAMICIA G; BADANO N; MAGLIOCO A; COSTA H; PIAZZON I; NEPOMNASCHY I

Congreso; First French-Argentine Immunology Congress; 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 de*cient 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 signi*cant 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 di´erences 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- were found in cultures of LN CTSLnkt cells using both RT-PCR and ELISA assays