Kidney Week

Abstract: TH-OR018

Organelle-Specific Oxidant Stress and CORE Disruption Mediate Proximal Tubule Cell Injury During Gentamicin Exposure

Session Information

• AKI: Mechanisms - Injury and Repair
  November 07, 2019 | Location: Salon A/B, Walter E. Washington Convention Center
  Abstract Time: 05:54 PM - 06:06 PM

Category: Acute Kidney Injury

• 103 AKI: Mechanisms

Authors

• Igwebuike, Chinaemere, Boston Univerisity School of Medicine , Boston, Massachusetts, United States

Chinaemere Igwebuike



Chinaemere Igwebuike is an MD/PhD candidate in the Molecular and Translational Medicine program at the Boston University School of Medicine. Chinaemere’s thesis work describes novel mechanisms of proximal tubule cell injury and identifies targeted therapeutic interventions. Specifically, he examines how the Cross-Organelle Stress Response (CORE) dysfunction may serve as an early therapeutic target. Chinaemere is an NIDDK F30-funded researcher and his thesis project encompasses clinical research identifying risk factors for acute kidney injury in socioeconomically disadvantaged patient populations.

• Pimentel, David, Boston Univerisity School of Medicine , Boston, Massachusetts, United States

David Pimentel,

• Feng, Hui, Boston Univerisity School of Medicine , Boston, Massachusetts, United States

Hui Feng,

• Campbell, Joshua, Boston Univerisity School of Medicine , Boston, Massachusetts, United States

Joshua Campbell,

• Sherman, Michael, Ariel University, Jerusalem, Israel

Michael Sherman,

• Yaglom, Julia, Ariel University, Jerusalem, Israel

Julia Yaglom,

• Wang, Zhiyong, Boston Medical Center, Boston, Massachusetts, United States

Zhiyong Wang,

• Havasi, Andrea, Boston Medical Center, Boston, Massachusetts, United States

Andrea Havasi,

• Borkan, Steven C., Boston Medical Center, Boston, Massachusetts, United States

Steven C. Borkan,

Background

The Cross-Organelle Stress Response (CORE) is an adaptive mechanism that maintains mitochondrial and endoplasmic reticulum (ER) proteostasis. We hypothesized that gentamicin causes nephrotoxic acute kidney injury (AKI) by causing both mitochondrial specific oxidative stress and fragmentation resulting in CORE disruption before activating the lethal unfolded protein response (UPR).

Methods

Mito HyPer, a mitochondrial-specific H₂O₂ probe, was used to detect early mitochondrial ROS accumulation in human proximal tubule epithelial cells (HK2) during gentamicin exposure. Mitochondria and ER were stained with MitoTracker and ER Tracker, respectively, and time course experiments were performed using a Nikon Super Resolution microscope. Mitochondrial-ER disassociation, mitochondrial morphology and immunoblots of CORE-associated mitochondrial pro-fission proteins (Total DRP1 and pDRP1) were used as surrogates of CORE function. Misfolded protein stains (Thioflavin T), protein ubiquitination, and immunoblots for whole cell oxidative stress (4HNE) were measured to assess proteotoxicity. The efficacy of preserving CORE on protein misfolding, lethal UPR activation (CHOP), and cell survival was assessed using geranylgeranylacetone (GGA), a protein chaperone inducer, prior to the introduction of gentamicin.

Results

Gentamicin exposure caused characteristic features of disrupted CORE, including mitochondrial-specific H₂O₂ accumulation, DRP-1 activation and organelle fragmentation, followed by mitochondrial-ER dissociation. Importantly, CORE disruption occurred before detectable changes in whole cell oxidative stress, protein ubiquitination, protein misfolding or lethal UPR activation (CHOP) were observed. GGA significantly decreased mitochondrial-specific oxidative stress, prevented fragmentation, preserved mitochondrial-ER association and ameliorated lethal UPR activation.

Conclusion

Gentamicin exposure causes early mitochondrial H₂O₂ accumulation and disrupts the CORE. These untoward events contribute to gentamicin-induced proteotoxicity and lethal UPR activation. GGA preserves the CORE and decreases subsequent lethal UPR activation that contributes to the proximal tubule cell injury caused by gentamicin.

Funding

• NIDDK Support