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Abstract: PO0376

Pulsed Ultrasound Reduces Oxidative Stress-Induced Disruption of Epithelial Barrier in Sepsis-AKI

Session Information

Category: Acute Kidney Injury

  • 103 AKI: Mechanisms

Authors

  • Zheng, Shuqiu, University of Virginia School of Medicine, Charlottesville, Virginia, United States
  • Rosin, Diane L., University of Virginia School of Medicine, Charlottesville, Virginia, United States
  • Yao, Junlan, University of Virginia School of Medicine, Charlottesville, Virginia, United States
  • Poudel, Nabin, University of Virginia School of Medicine, Charlottesville, Virginia, United States
  • Tanaka, Shinji, University of Virginia School of Medicine, Charlottesville, Virginia, United States
  • Nash, William, University of Virginia School of Medicine, Charlottesville, Virginia, United States
  • Okusa, Mark D., University of Virginia School of Medicine, Charlottesville, Virginia, United States
Background

Oxidative stress disrupts epithelial junctions leading to increased paracellular permeability and kidney dysfunction. We previously showed that pulsed ultrasound (pUS) reduced inflammation and kidney injury. We hypothesized that pUS mitigates renal injury by maintaining epithelial tight junctions. Here, we utilized lipopolysaccharide (LPS) induced sepsis to create acute kidney injury (S-AKI) in a mouse model and RAW 264.7 cells to investigate the effects of pUS on the epithelial tight junction barrier and renal macrophages.

Methods

C57/BL/6 mice received pUS 24 hrs before LPS treatment. The parameters of pUS therapy followed the protocol previously published by us (PMID: 23907510). Following pUS treatment, mice received a single injection of LPS (5 mg/kg, ip). Animals were euthanized at increasing time intervals for measurement of mRNA expression and kidney imaging. Kidney histopathological changes were observed by using PAS staining. Co-staining with TUNEL and cleaved caspase-3 was used to assess kidney injury. For in-vitro assays, RAW cells were seeded onto 4-well plates and incubated for 24 hrs at a density of 5 × 105 per well. Cells were treated with LPS (100 ng/mL) in serum-free DMEM for 2 hrs.

Results

LPS induced kidney injury and apoptosis, as observed by PAS and TUNEL staining, was attenuated by pUS. Co-staining with PSD95 (postsynaptic scaffolding density protein 95) and ZO-1 (zonula occludens-1) showed both were expressed in kidney. LPS also induced a significant loss of PSD95 accompanied by a reduced mRNA expression of nuclear factor erythroid 2-related factor 2 (NRF2) and activated macrophages. The structural changes, extent of loss of PSD95 and NRF2, as well as macrophage infiltrate were all partially reversed by prior pUS treatment. In cultured RAW cells, pUS upregulated the expression of NRF2 and heme oxygenase-1(HO-1), and attenuated CD68-positive macrophage signals.

Conclusion

pUS protected kidneys from LPS-induced S-AKI by preserving antioxidant NRF2 expression and attenuating oxidative stress-induced disruption of epithelial tight junctions.

Funding

  • NIDDK Support