Kidney Week

Abstract: SA-PO791

Increased Renal Inflammation in Drd5 Knockout Mice Is Associated With Peroxiredoxin-4 Dysfunction

Session Information

• Hypertension and CVD: Mechanisms
  November 05, 2022 | Location: Exhibit Hall, Orange County Convention Center‚ West Building
  Abstract Time: 10:00 AM - 12:00 PM

Category: Hypertension and CVD

• 1503 Hypertension and CVD: Mechanisms

Authors

• Amatya, Bibhas, The George Washington University School of Medicine and Health Sciences, Washington, District of Columbia, United States

Bibhas Amatya,

• Polzin, Jacob Quentin Mullins, The George Washington University School of Medicine and Health Sciences, Washington, District of Columbia, United States

Jacob Quentin Mullins Polzin,

• Asico, Laureano D., The George Washington University School of Medicine and Health Sciences, Washington, District of Columbia, United States

Laureano D. Asico,

• Armando, Ines, The George Washington University School of Medicine and Health Sciences, Washington, District of Columbia, United States

Ines Armando,

• Felder, Robin Allen, University of Virginia Health Sciences Center, Charlottesville, Virginia, United States

Robin Allen Felder,

• Jose, Pedro A., The George Washington University School of Medicine and Health Sciences, Washington, District of Columbia, United States

Pedro A. Jose,

• Lee, Hewang, The George Washington University School of Medicine and Health Sciences, Washington, District of Columbia, United States

Hewang Lee,

Background

The dopamine D5 receptor (D₅R) suppresses inflammation in the central nervous system. However, its role in the regulation of inflammation associated with oxidative stress and hypertension in the kidney is not known.

Methods

Protein expression was determined by immunoblotting in D₅R-expressing cells, treated with pharmacological reagents or transfected with mock or specific siRNA to silence specific genes. Protein-protein interaction was determined by co-immunoprecipitation and co-localization analysis of immunofluorescence images. Drd5^-/- mice were generated, as previously reported.

Results

In D₅R-HEK 293 cells, fenoldopam (FEN, 25 nM/12 hr, n=4), a D₁-like receptor agonist, increased the expression (158.1±11.9% vs 100.0±6.8% basal level, n=6) of peroxiredoxin-4 (PRDX4), an endoplasmic reticulum-localized protein. By contrast, D₅R protein was decreased in PRDX4 siRNA-treated D₅R-HEK293 (58.8±6.7% vs 100.0±9.6% basal level, n=4) and -human renal proximal tubule cells (hRPTCs) (60.4±5.8%, n=4). PRDX4 protein was also decreased in the kidney cortices of Ddr5^-/- mice (Ddr5^+/+: 100±18%, n=5; D₅R^-/-: 69±14%, n=4; P<0.05). FEN increased the co-immunoprecipitation of D₅R and PRDX4 and their colocalization, particularly in the endoplasmic reticulum. Moreover, silencing PRDX4, by its specific siRNA, increased reactive oxygen species (ROS) production and impaired the inhibitory effect of FEN on ROS production in hRPTCs. In D₅R-HEK 293 cells, siRNA silencing of PRDX4 also increased the production of tumor necrosis factor (TNF) and interleukin (IL)-1b. The increase in TNF protein in D₅R-HEK293 cells with silenced PRDX4 was attenuated by tempol, a superoxide dismutase mimetic, indicating that oxidative stress and downstream inflammation are associated with PRDX4 deficiency and consistent with the increase in NLRP3 inflammasome with PRDX4 silencing (scrambled siRNA: 100±5.0%, PRDX4 siRNA: 150.6±6.0%). The protein expressions of TNF (Ddr5^+/+: 100±11%, n=4; D₅R^-/-: 179±16%, n=4; P<0.05) and IL-1β (Ddr5^+/+: 100±13%, n=4; D₅R^-/-: 156±14%, n=4; P<0.05) were also increased in the kidney cortices of Drd5^-/- mice, relative to Drd5^+/+ littermates.

Conclusion

The increase in renal inflammation in Drd5^-/- mice is due, in part, to oxidative stress, caused by the impaired interaction between D₅R with PRDX4.

Funding

• NIDDK Support