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Abstract: FR-PO200

Podocyte Nox5 Expression Increases Microparticle Formation and Oxidative Stress in Mice

Session Information

Category: Diabetic Kidney Disease

  • 601 Diabetic Kidney Disease: Basic

Authors

  • Battaion, Hannah L., Ottawa Hospital Research Institute, Ottawa, Ontario, Canada
  • Holterman, Chet E., Ottawa Hospital Research Institute, Ottawa, Ontario, Canada
  • Burger, Dylan, Kidney Research Centre, Ottawa, Ontario, Canada
  • Kennedy, Chris R., Kidney Research Centre, Ottawa, Ontario, Canada
Background

Urinary microparticles (MPs) released from podocytes are predictive of injury associated with diabetic kidney disease (DKD) prior to albuminuria. Oxidative stress due to reactive oxygen species (ROS) overproduction is one factor that contributes to MP formation. Podocyte NADPH oxidase 5 (Nox5) produces superoxide and is upregulated in human DKD. Transgenic mice expressing podocyte-specific Nox5 (Nox5Pod+) show increased blood pressure, glomerular basement membrane thickening and albuminuria. However, whether podocytes expressing Nox5 produce more MPs under conditions of stress associated with diabetes and hypertension, and whether these microparticles contain Nox5 and are capable of producing reactive oxygen species remains unknown.

Methods

For in vitro studies, Nox5 was activated in conditionally immortalized human podocytes through exposure to ionomycin for 24 hours. MPs released into the media were isolated and quantified by nanoparticle tracking analysis. Nox5 content and ROS production from isolated MPs were assessed by immunoblotting and DHE/HPLC, respectively. For mouse studies, Nox5Pod+ mice and nontransgenic littermate controls were either implanted with angiotensin II-containing miniosmotic pumps (400mg/kg/day) or sham operated.Systolic blood pressures were determined by tail cuff plethysmography.Urinary MPs were quantified once per week for 5 weeks using 24 hour urine collections.

Results

MP formation was increased two-fold in cultured human podocytes when stimulated with Ionomycin compared to unstimulated controls. Isolated podocyte MPs expressed Nox5 and demonstrated increased ROS formation compared to unstimulated controls.Urinary MP formation was increased in Nox5Pod+ mice compared to controls. Angiotensin II administration increased systolic blood pressure in both Nox5Pod+ and nontransgenic mice.However, MP formation was further increased following angiotensin administration.

Conclusion

Nox5-expressing podocytes release MPs. These MPs contain Nox5 and are capable of producing ROS.Podocyte Nox5 expression in mice show enhanced urinary MP formation in the context of high blood pressure. Thus, Nox5 may be a contributing factor in the onset of damage in DKD and/or hypertension.

Funding

  • Government Support - Non-U.S.