Abstract: FR-PO217

D-Serine, a Novel Uremic Toxin, Induces Senescence in Human Renal Tubular Cells via GCN2 Activation

Session Information

Category: Cell Biology

  • 202 Apoptosis, Proliferation, Autophagy, Cell Senescence, Cell Transformation

Authors

  • Okada, Akira, The University of Tokyo Graduate School of Medicine, Tokyo, Japan
  • Jao, Tzu-Ming, The University of Tokyo Graduate School of Medicine, Tokyo, Japan
  • Maekawa, Hiroshi, The University of Tokyo Graduate School of Medicine, Tokyo, Japan
  • Ishimoto, Yu, The University of Tokyo Graduate School of Medicine, Tokyo, Japan
  • Kawakami, Takahisa, The University of Tokyo Graduate School of Medicine, Tokyo, Japan
  • Nangaku, Masaomi, The University of Tokyo Graduate School of Medicine, Tokyo, Japan
  • Inagi, Reiko, The University of Tokyo Graduate School of Medicine, Tokyo, Japan
Background

A recent metabolomic analysis revealed the change in profile of metabolites, including uremic toxins, in plasma or urine from CKD patients. Accumulation of D-serine, an enantiomer of L-serine, in plasma is associated with faster progression of renal dysfunction in CKD patients. We investigated whether chirality of the amino acid plays a crucial role in the pathogenesis of CKD.

Methods

To address the effect of D-serine on tubules, human proximal tubular cell line, HK-2, and primary culture of human renal tubular cells, NHREC, were treated with D- or L-serine for 48 hr and the cell damages were evaluated by cell proliferation (MTS assay and cell count), cell cycle status (PI and Phospho-Histone H3 staining), senescence (p21, p16, γH2AX, and SA-βG), senescence-associated secretory phenotype [SASP: pro-inflammatory cytokines (IL-6 and IL-8)] and apoptosis (Annexin V staining and caspase 3/7 activity). To find out the molecular mechanism of the cell damages by D-serine, we assessed the status of amino acid-mediated signaling (integrated stress response: GCN2, ATF4, and CHOP) and L-serine synthesis pathway (PHGDH and PSAT1), To confirm signal transduction, siRNAs of integrated stress response molecules were used.

Results

D-serine, but not L-serine, markedly induced cellular senescence and apoptosis both in HK-2 and NHREC. Such tubular damage by D-serine was accompanied by G2/M cell cycle arrest and induction of SASP, including pro-inflammatory factors, contributing to tubulointerstitial fibrosis. Importantly, we found that integrated stress response mediated by GCN2-ATF4-CHOP pathway played a central role in D-serine-induced cell toxicity: knockdown of GCN2 ameliorated D-serine-induced tubular cell senescence, suggesting CKD progression and kidney aging by D-serine. Furthermore, D-serine upregulated the L-serine synthesis pathway, possibly as a counteracting mechanism, and D-serine-induced tubular toxicity is counteracted by L-serine, suggest that the proportion of D-/L-serine is critical for D-serine toxicity to tubular cells.

Conclusion

This study unveils a pathogenic role of the chiral amino acid and molecular mechanisms underlying D-serine-induced tubular damage, such as senescence with SASP, in CKD pathogenesis.

Funding

  • Government Support - Non-U.S.