Intrarenal crystal formation activates the Nlrp3 inflammasome in myeloid cells and triggers a profound inflammatory response. Here, we studied whether a specific inhibitor of the Nlrp3 inflammasome, CP-456,773, can prevent kidney fibrosis in a murine model of crystal nephropathy induced by diets rich in oxalate or adenine. Inflammasome activation in renal dendritic cells and the resulting interleukin (IL)-1β and IL-18 production were markedly reduced by CP-456,773 treatment both ex vivo and in vivo. We directly visualized intrarenal inflammasome activation and its inhibition by CP-456,773 in vivo by adoptive transfer of bone marrow cells transduced with interleukin-1β-Gaussia luciferase, a proteolytic luciferase-based reporter for inflammasome activation, into irradiated mice. CP-456,773 treatment strongly attenuated kidney fibrosis when given early in the genesis of crystal nephropathy, but was unable to reverse established crystal-induced fibrosis. The urinary IL-18 concentration appeared to be a useful noninvasive biomarker for renal inflammasome activation. Finally, NLRP3 inhibition did not compromise adaptive immune responses as previously reported for the global inhibition of IL-1 signaling. Thus, early NLRP3 inhibition by CP-456,773 may be an effective treatment for crystal nephropathy. Use of iGLuc transfected cells introduces a novel imaging technique for inflammasome activation in mice.

Regulatory T cells (Tregs) maintain self-tolerance and prevent autoimmunity by controlling autoreactive T cells. We recently demonstrated in vivo that Tregs can directly suppress auto-reactive B cells via programmed death ligand 1 (PD-L1) that ligated PD-1 on B cells and caused them to undergo apoptosis. Here, we asked whether this mechanism is utilized by thymus-derived natural Tregs and/or by peripheral lymphoid tissue-induced Tregs. We first demonstrated that antigen-specific PD-L1-expressing Tregs were induced in the draining lymph node of autoantigen-expressing tissue and characterized them by their lack of the transcription factor Helios and of the surface marker Neuropilin-1 (Nrp-1). Next, we established an in vitro co-culture system to study the interaction between B cells and Treg subsets under controlled conditions. We found that Nrp Treg, but not Nrp Treg suppressed autoreactive B cells, whereas both were able to suppress T-helper cells. Such suppression was antigen-specific and was facilitated by PD-L1/PD-1-induced apoptosis. Furthermore, it required physical cell contact and was MHC II-restricted, providing an explanation for the antigen-specificity of peripherally-induced Tregs. These findings identify a role for peripherally induced Helios Nrp-1 inducible Treg in controlling peripheral B-cell tolerance against tissue auto-antigens.

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