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Abstract: SA-OR14

Serial Intravital Imaging of Ischemia-Reperfusion Injury Reveals the Dynamics of Tubular Injury and Repair

Session Information

Category: Acute Kidney Injury

  • 103 AKI: Mechanisms

Authors

  • Bordoni, Luca, Aarhus Universitet, Aarhus, Midtjylland, Denmark
  • Pohl, Layla, Aarhus Universitet, Aarhus, Midtjylland, Denmark
  • Kristensen, Anders Meldgaard, Aarhus Universitet, Aarhus, Midtjylland, Denmark
  • Kidmose, Hanne, Aarhus Universitet, Aarhus, Midtjylland, Denmark
  • Sardella, Donato, Aarhus Universitet, Aarhus, Midtjylland, Denmark
  • Schiessl, Ina M., Aarhus Universitet, Aarhus, Midtjylland, Denmark

Group or Team Name

  • Laboratory of Renal Cell Remodeling and Regeneration (The Schiessl-lab)
Background

The kidney has a remarkable capacity to recover from acute kidney injury (AKI), but the involved dynamics of cellular damage and repair are incompletely understood. In this study, we investigated ischemia-reperfusion injury (IRI)-induced proximal tubule (PT) cell death and proliferation of the same renal cells over time using serial intravital 2-photon microscopy (2PM).

Methods

We performed 21 minutes IRI of the left kidney followed by abdominal imaging window implantation for serial imaging of the same tissue at 1 h and day 1, 2, 3, 4, 7, 14 and 21 after IRI. CycB1-GFP reporter mice identified proliferating cells by GFP-expression in S-G2-M cell cycle-stages.

Results

Necrotic tubular cell death, as detected by Propidium Iodide (PI)-staining, was primarily observed 1 h post IRI and mostly affecting PTs with 10.1±4.1 %, 12.3±3.8% and 1.9±.8% PI-positive nuclei per S1, S2 and distal tubule (DT)/collecting duct (CD) segments (Mean±SEM, n=8 each). From day 1, injured PTs shedded brushborder contents, which correlated with epithelial flattening and onset of proliferation (p=.002, r2=.39). Tubular proliferation started day 1, peaked day 3 and was highest in S2 segments with 2.7±2%, 12.4±8.7% and 0.3±.9% (Mean±SEM, n=8 each) of GFP-positive nuclei per S1, S2 and DT/CD segments (p=.02. for S2 vs. S1 and p>.001 for S2 vs. DT/CD). While in S1 segments proliferation derived mainly from surviving cells in immediate proximity to PI-positive cells, in S2 segments also cells further distant from injured sites proliferated. We observed shedded cytosolic content from injured PT regions flowing downstream into previously PI-negative PTs, which spatially coincided with their proliferation one day after the appearance of shedded material in the lumen. By day 4, several PT segments revealed severe cast formation and epithelial vacuolization with nuclear karyolysis. 75% of the vacuolized tubule population reached full recovery before day 14 post IRI, while the remaining 25% failed to recover, resulting in nephron loss.

Conclusion

This is the first study to track IRI-induced injury and regeneration in the same renal cells over time. Our data uniquely links initial tissue damage to regenerative capacity of the renal PT in AKI and suggests distinct mechanisms for initiation of PT proliferation in S1 and S2 segments.

Funding

  • Private Foundation Support