Upregulating NRF2 Is a Critical Regulatory Mechanism for the Protective Effect of Ultrasound to Mitigate Sepsis-Associated AKI

Background: Sepsis-induced oxidative stress and dysfunction of innate immune response have emerged as key players in the pathogenesis of sepsis-associated acute kidney injury (S-AKI). Nuclear factor-erythroid-derived 2-related factor 2 (NRF2), a key oxidative stress regulator, has been implicated to play an important role in T cell-specific augmentation of Nrf2 to mitigate oxidative stress. Our previous studies indicated that pulsed ultrasound (pUS) can reduce inflammation and acute kidney injury in mice. We hypothesis that pUS mediated protection is associated with enhanced the expression of Nrf2 and reduced immune system dysfunction. In this study, we utilized LPS-induced S-AKI in normal and Nrf2-/- mice, and we used RAW264.7 cells to investigate the effects of pUS on LPS-induced kidney injury, macrophage filtration and NRF2. In addition, we also investigated whether pUS protection occurs through enhanced NRF2 expression and disturbed CD4+ T cells immune function in human Jurkat T cells. Methods: C57/BL/6 mice received pUS 24 hours before LPS (5 mg/kg, ip) treatment. The parameters of pUS therapy followed the protocol we previously published. In vitro studies were performed using RAW cells and Jurkat T-cell lines, which were cultured and stimulated with LPS (100ng/ml) for 6h. NRF2 protein and mRNA expression was measured by immunofluorescence and RT-PCR respectively. Kidney injury was assessed by Kim1, cleaved-caspase-3 and plasma creatinine assay. Iba1, F4/80 and CD4+ T cells were used to evaluate immune cells/macrophage infiltration. Results: LPS produced AKI and macrophage/lymphocyte filtration in WT mice with a dramatic decline of NRF2. Furthermore, pUS-treated mice with high Nrf2 expression had fewer Iba1+ and F4/80+ macrophage infiltration in the kidney and CD4+ T cells in Jurkat T cells and attenuated sepsis-induced AKI. In contrast, LPS stimulus induced greater infiltration of microphages, as well as more early time and more severe renal injury in Nrf2-/- mice compared with litter mate control mice. pUS attenuated S-AKI. Conclusions: These results suggest that upregulating Nrf2 antioxidant defenses in kidney and in T cells are essential for ultrasound to attenuate oxidative stress-induced AKI. Our results reveal a novel mechanism for pUS protection from kidney injury during S-AKI. Funding: NIDDK Support