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Kidney Week

Abstract: SA-PO1192

Urea Suppresses the Alternative Activation of Murine Macrophages

Session Information

  • CKD: Mechanisms - 3
    October 26, 2024 | Location: Exhibit Hall, Convention Center
    Abstract Time: 10:00 AM - 12:00 PM

Category: CKD (Non-Dialysis)

  • 2303 CKD (Non-Dialysis): Mechanisms

Authors

  • Chae, Seung Yun, The Catholic University of Korea Seoul St Mary's Hospital, Seoul, Korea (the Republic of)
  • Min, Kyoung Il, The Catholic University of Korea Seoul St Mary's Hospital, Seoul, Korea (the Republic of)
  • Kim, You-Me, Korea Advanced Institute of Science and Technology, Daejeon, Korea (the Republic of)
Background

Although urea is generally considered harmless, it has been documented to obstruct the intracellular transport of arginine in vascular endothelial cells and inhibit glycolysis in pancreatic beta cells. This study aims to determine whether elevated extracellular urea concentrations impact macrophage functions.

Methods

Peritoneal macrophages and bone marrow-derived macrophages (BMDMs) were exposed to LPS and IL-4 to induce M1 and M2 polarization, respectively, in the presence of varying urea concentrations. Flow cytometry was utilized to examine the expression of markers of macrophage polarization including arginase-1 and iNOS. Additionally, M2 marker gene expression was analyzed using qPCR. Bulk RNA sequencing was performed to elucidate the transcriptomic changes induced by urea in BMDMs. The Seahorse real-time cell metabolic analysis was conducted to measure the oxygen consumption rate (OCR) and extracellular acidification rate (ECAR) in IL-4-treated BMDMs.

Results

Our study discovered that physiologically high urea concentrations reduce IL-4-induced upregulation of arginase-1 in murine macrophages in a dose-dependent manner at both mRNA and protein levels. Urea also significantly decreased the expression of other M2 macrophage markers, such as Fizz1 and Ym1. However, urea had minimal impact on LPS-induced M1 polarization. RNA sequencing of M2-polarized BMDMs indicated that urea downregulates genes associated with mTOR signaling, glycolysis, and oxidative phosphorylation. Correspondingly, urea treatment reduced phosphorylation of S6 protein and AKT, targets of mTORC1 and mTORC2, respectively. Additionally, ECAR and OCR were lower in urea-treated M2 BMDMs. The attenuation of M2 polarization by urea was not evident in the presence of Torin, a pan mTOR inhibitor, indicating that urea's inhibitory effect is mediated through downregulation of mTOR signaling.

Conclusion

Based on these findings, we propose that, contrary to the traditional view of urea as an inert molecule, urea suppresses M2 polarization in murine macrophages, likely through the attenuation of mTOR signaling.

Flow cytometry analysis of M2 marker expression in BMDMs