Kidney Week

Abstract: SA-PO566

Kidney Injury Under Oxidative Stress Release CD36 and CD47 Microparticles

Session Information

• AKI: Other Mechanisms and Cell Cultures
  October 27, 2018 | Location: Exhibit Hall, San Diego Convention Center
  Abstract Time: 10:00 AM - 12:00 PM

Category: Acute Kidney Injury

• 103 AKI: Mechanisms

Authors

• Campos, Begoña, University of Cincinnati, Cincinnati, Ohio, United States

Begoña Campos,

• Bockhorst, Samuel Paul, University of Cincinnati, Cincinnati, Ohio, United States

Samuel Paul Bockhorst,

• Saum, Keith Louis, University of Cincinnati, Cincinnati, Ohio, United States

Keith Louis Saum,

• Thakar, Charuhas V., University of Cincinnati, Cincinnati, Ohio, United States

Charuhas V. Thakar,

Group or Team Name

• Kidney Injury Translational Research Group

Background

Kidney epithelial cell damage is the major cause of acute kidney injury (AKI). Thrombospondin 1 (TSP-1) is released by renal epithelial cells and mediates kidney damage in murine ischemia reperfusion injury (IRI). TSP-1 exerts its inflammatory modulating effects through signaling of CD36 (pro-inflammatory) and CD47 (anti-inflammatory) ligands. Given that CD36 reduces angiogenesis, and stimulates VEGF induced migration and apoptosis, whereas CD47 stimulate angiogenesis, these ligands may play a role in epithelial to endothelial cross-talk. We hypothesized that in both in vivo murine models of IRI and in vitro models of oxidative stress to human renal epithelial cells (h-RPTEC), CD36 and CD47 proteins will be released as MP significantly when compared to control.

Methods

h-RPTEC line was used and oxidative stress (induced by adding 0.03 mM of H₂O₂ for 1hr) was evaluated for MP release and apoptosis compared to controls. IRI was performed in C57BL/6 mice by renal pedicle clamp for 30 minutes followed by reperfusion compared to sham. Animals were euthanized at 48 hrs. Histology was performed by standard techniques to evaluate kidney damage. Citrate poor plasma was collected at 0, 12, 48 hrs to measure urea (BUN). MP were measured with flow cytometry and were reported at 10⁵ /ml and analyzed by flow jo software. Unpaired t test Welch’s correction were used for comparisons.

Results

h-RPTEC contained CD36 and CD47 receptors in the cellular membrane, confirmed by immunohistochemistry. Upon exposure to H₂O_2, cell viability study showed 25% apoptotic, 15% in necrotic and 60% viable. A a significant increase of release of MP was observed for CD36 vs control (2.7 X10⁵ MP/ml vs. 0.73 X10⁵ MP/ml; p=0.001); and for CD47 vs control (31.84 10⁵ MP/ml vs 2.51 10⁵ MP/ml; p=0.001). For in vivo studies, compared to sham, IRI mice showed that MP containing CD36 were released at 12 hr and remained elevated at 48 hr. In contrast, CD47 showed an increase at 12 hrs and returned towards baseline by 48 hr. Analysis of percent expression showed that CD36 expression increased with time compared with CD47.

Conclusion

In murine and in vitro models of oxidative stress CD36 and CD47 are released as MP. Given the role of TSP-1 and other thrombomodulins in AKI, MP may serve as ligands to both potentiate epithelial cell injury as well as mediate cross-talk between epithelium and endothelium.

Funding

• Clinical Revenue Support