Background: A highfrequency of regulatory B (Breg) cells, generally transitional B cells (Btrans), has been associated with long-term kidney allograft survival and operational tolerance, while circulating follicular helper T cells (cTfh) correlate with graft rejection. We aimed to understand the interplay between these cell subsets and to determine their association with renal graft outcome.
Methods: Btrans (CD19+CD24+CD38+) and IL10+Breg cells (CD19+CD24highCD38high cells producing IL10 after CpG+CD40L ex vivo stimulation), as well as cTfh (CD4+CXCR5+T cells) have been analysed, pre- and post-transplantation, in a prospective cohort of 200 kidney transplant recipients and in healthy volunteers (HV). 
Results: End-stage renal disease patients had higher frequencies of Btrans and IL10+Breg cells compared to HV(20% vs 10% and 1.5% vs 0.8%, p<0.001 and p<0.01, respectively). Pre-transplantation frequencies of Btrans and IL10+Breg cells were significantly higher in patients later rejecting than in patients remaining stable (26% vs 20%, p<0.05, and 2.3% vs 1.5%, p<0.01, respectively).In the multivariate analysis, pre-transplantation %IL10+ Breg above the median predicted rejection and graft failure independently from age, de novoanti-HLA antibodies and delayed graft function (HR=3.98 and 13.46 respectively, both p=0.02). IL10+Bregdecreased during the 1-year post-transplant follow-up, and a larger pre- to 7 days post-transplant reduction of %IL10+ Breg was also an independent predictor of rejection and graft failure (HR=3.63 and 5.86). Finally, IL10+ Breg correlated with cTfh pre-transplantation (r=0.28, p<0.01). The cTfh/IL10+Breg ratio increased after transplantation, and this augmentation preceded and was significantly higher in rejectors vs stable recipients (p<0.05). 
Conclusions: Evaluation of pre-transplant IL10+Breg cells and the regular monitoring of the cTfh/IL10+ Breg ratio may be useful to assess post-transplant risk. Additionally, these observations suggest the need to develop therapeutic strategies aimed at depleting Tfh while preserving regulatory B cells in transplanted patients.