Evaluation of Neutrophil Dynamics Change by Protective Effect of Tadalafil After Renal Ischemia/Reperfusion Using In Vivo Real-time Imaging.

BACKGROUND Neutrophils play a major role in ischemia/reperfusion injury (IRI) in renal transplantation and acute kidney injury. However, it has been difficult to observe changes in neutrophil dynamics over time in living mice kidney. We investigate neutrophil dynamics in IRI in living mice using novel in vivo multiphoton microscope imaging techniques and characterize the renoprotective effects of a selective phosphodiesterase (PDE) 5 inhibitor, tadalafil. METHODS Wild-type (WT) and eNOS knockout (eNOS-KO) mice, a model of endothelial dysfunction, were used to establish in vivo real-time imaging in living mouse kidneys. Neutrophils were labeled green with Ly-6G monoclonal antibody, and plasma flow was labeled red with bovine serum albumin. Tadalafil was administered orally 1 h before surgery. Both kidney pedicles were reperfused after 37° warm ischemia for 45 min. RESULTS Our novel approach revealed that neutrophils were trapped in glomerulus within a few minutes after reperfusion. They gradually increased over time and Infiltrated neutrophils were observed in the tubular lumen and peritubular capillary. The neutrophils were clearly visualized rolling on peritubular capillary plexus at 3 μm/min. The administration of tadalafil significantly reduced neutrophil influx into the glomerulus in both WT and eNOS-KO mice. Reduced neutrophil infiltration in tadalafil groups, which was confirmed by flow cytometry, resulted in histopathologically decreased tubular injury. The expression of VCAM-1 and KIM-1 was partially prevented by tadalafil. CONCLUSIONS Use of a novel technique contributed to elucidation of neutrophil dynamics after reperfusion. Tadalafil has a potential for inhibiting neutrophil infiltration in renal IRI.Supplemental Visual Abstract; http://links.lww.com/TP/C223.