Peptidyl arginine deiminase-4 activation exacerbates kidney ischemia-reperfusion injury.

Peptidyl arginine deiminase (PAD)4 is a nuclear enzyme that catalyzes the posttranslational conversion of arginine residues to citrulline. Posttranslational protein citrullination has been implicated in several inflammatory autoimmune diseases, including rheumatoid arthritis, colitis, and multiple sclerosis. Here, we tested the hypothesis that PAD4 contributes to ischemic acute kidney injury (AKI) by exacerbating the inflammatory response after renal ischemia-reperfusion (I/R). Renal I/R injury in mice increased PAD4 activity as well as PAD4 expression in the mouse kidney. After 30 min of renal I/R, vehicle-treated mice developed severe AKI with large increases in plasma creatinine. In contrast, mice pretreated with PAD4 inhibitors (2-chloroamidine or streptonigrin) had significantly reduced renal I/R injury. Further supporting a critical role for PAD4 in generating ischemic AKI, mice pretreated with recombinant human PAD4 (rPAD4) protein and subjected to mild (20 min) renal I/R developed exacerbated ischemic AKI. Consistent with the hypothesis that PAD4 regulates renal tubular inflammation after I/R, mice treated with a PAD4 inhibitor had significantly reduced renal neutrophil chemotactic cytokine (macrophage inflammatory protein-2 and keratinocyte-derived cytokine) expression and had decreased neutrophil infiltration. Furthermore, mice treated with rPAD4 had significantly increased renal tubular macrophage inflammatory protein-2 and keratinocyte-derived cytokine expression as well as increased neutrophil infiltration and necrosis. Finally, cultured mouse kidney proximal tubules treated with rPAD4 had significantly increased proinflammatory chemokine expression compared with vehicle-treated cells. Taken together, our results suggest that PAD4 plays a critical role in renal I/R injury by increasing renal tubular inflammatory responses and neutrophil infiltration after renal I/R.

Ham A ，Rabadi M ，Kim M ，Brown KM ，Ma Z ，D'Agati V ，Lee HT ... -  《-》

Peptidyl arginine deiminase-4-deficient mice are protected against kidney and liver injury after renal ischemia and reperfusion.

We previously demonstrated that renal peptidyl arginine deiminase-4 (PAD4) is induced after renal ischemia and reperfusion (I/R) injury and exacerbates acute kidney injury (AKI) by increasing the renal tubular inflammatory response. Here, we tested whether genetic ablation of PAD4 attenuates renal injury and inflammation after I/R in mice. After renal I/R, PAD4 wild-type mice develop severe AKI with large increases in plasma creatinine, neutrophil infiltration, as well as significant renal tubular necrosis, apoptosis, and proinflammatory cytokine generation. In contrast, PAD4-deficient mice are protected against ischemic AKI with reduced real tubular neutrophil infiltration, renal tubular necrosis, and apoptosis. In addition, hepatic injury and inflammation observed in PAD4 wild-type mice after renal I/R are significantly attenuated in PAD4-deficient mice. We also show that increased renal tubular PAD4 expression after renal I/R is associated with translocation of PAD4 from the nucleus to the cytosol. Consistent with PAD4 cytosolic translocation, we show increased renal tubular cytosolic peptidyl-citrullination after ischemic AKI. Mechanistically, recombinant PAD4 treatment increased nuclear translocation of NF-κB in cultured human as well as murine proximal tubule cells that is inhibited by a PAD4 inhibitor (2-chloroamidine). Taken together, our studies further support the hypothesis that renal tubular PAD4 plays a critical role in renal I/R injury by increasing the renal tubular inflammatory response and neutrophil infiltration after renal I/R perhaps by interacting with the proinflammatory transcription factor NF-κB in the cytosol and promoting its nuclear translocation.

Rabadi M ，Kim M ，D'Agati V ，Lee HT ... -  《-》

Peptidyl arginine deiminase-4 exacerbates ischemic AKI by finding NEMO.

Rabadi MM ，Han SJ ，Kim M ，D'Agati V ，Lee HT ... -  《-》

Neutrophil peptidyl arginine deiminase-4 has a pivotal role in ischemia/reperfusion-induced acute kidney injury.

Ischemia/reperfusion is a common cause of acute kidney injury (AKI). However, mechanisms underlying the sudden loss in kidney function and tissue injury remain to be fully elucidated. Here, we investigated the role of peptidyl arginine deiminase-4 (PAD4), which converts arginine to citrulline and plays a role in epigenetic regulation and inflammation, in renal ischemia/reperfusion injury. PAD4 expression was highly induced in infiltrating leukocytes 24 hours following renal ischemia and reperfusion. This induction was accompanied by citrullination of histone H3 and formation of neutrophil extracellular traps in kidneys of wild-type mice. By contrast, PAD4-deficient mice did not form neutrophil extracellular traps, expressed lower levels of pro-inflammatory cytokines and were partially protected from renal ischemia/reperfusion-induced AKI. Furthermore, PAD4-deficient mice recovered kidney function 48 hours after ischemia/reperfusion, whereas kidney function in the wild-type mice progressively worsened. Administration of DNase I, which degrades neutrophil extracellular traps or the PAD-specific inhibitor YW3-56 before ischemia, partially prevented renal ischemia/reperfusion-induced AKI. Notably, transfer of neutrophils from wild-type, but not from PAD4-deficient mice, was sufficient to restore renal neutrophil extracellular trap formation and impair kidney function following renal ischemia/reperfusion. Thus, neutrophil PAD4 plays a pivotal role in renal ischemia/reperfusion-induced AKI.

Raup-Konsavage WM ，Wang Y ，Wang WW ，Feliers D ，Ruan H ，Reeves WB ... -  《-》

Divergent roles for kidney proximal tubule and granulocyte PAD4 in ischemic AKI.

Li H ，Han SJ ，Kim M ，Cho A ，Choi Y ，D'Agati V ，Lee HT ... -  《-》