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Kidney Week

ASN / Education & Meetings / Kidney Week /
Abstract: TH-PO0132

MANF Clears Mutant Uromodulin in Human Kidney Organoids of Autosomal Dominant Tubulointerstitial Kidney Disease

Session Information

Category: Genetic Diseases of the Kidneys

  • 1202 Genetic Diseases of the Kidneys: Non-Cystic (Complex and Non-Cystic Monogenic)

Authors

  • Gu, Chenjian, Washington University in St Louis School of Medicine, St. Louis, Missouri, United States
  • Fang, Yili, Washington University in St Louis School of Medicine, St. Louis, Missouri, United States
  • Wang, Yixuan, Washington University in St Louis, St. Louis, Missouri, United States
  • Tycksen, Eric, Washington University in St Louis School of Medicine, St. Louis, Missouri, United States
  • Kondepati, Gayathri, Washington University in St Louis, St. Louis, Missouri, United States
  • Li, Chuang, Washington University in St Louis School of Medicine, St. Louis, Missouri, United States
  • Kidd, Kendrah O., Wake Forest University, Winston-Salem, North Carolina, United States
  • Liu, Jun, Shanghai General Hospital, Shanghai, China
  • Urano, Fumihiko, Washington University in St Louis School of Medicine, St. Louis, Missouri, United States
  • Lindahl, Maria, Helsingin yliopisto, Helsinki, Uusimaa, Finland
  • Bleyer, Anthony J., Wake Forest University, Winston-Salem, North Carolina, United States
  • Singamaneni, Srikanth, Washington University in St Louis, St. Louis, Missouri, United States
  • Sun, Zhao, The University of Texas at Dallas, Richardson, Texas, United States
  • Chen, Ying Maggie, Washington University in St Louis School of Medicine, St. Louis, Missouri, United States
Background

Autosomal dominant tubulointerstitial kidney disease due to UMOD mutations (ADTKD-UMOD) is a leading monogenic cause of chronic kidney disease, and the p.H177-R185del accounts for 21% of cases in the US. While misfolded UMOD is known to induce endoplasmic reticulum (ER) stress and impair autophagy in thick ascending limb (TAL) cells, effective therapies remain unavailable. We aimed to investigate the therapeutic potential and mechanism of mesencephalic astrocyte-derived neurotrophic factor (MANF), a novel ER chaperone, using patient-derived ADTKD organoids.

Methods

Induced pluripotent stem cells (iPSCs) were derived from a heterozygous UMOD H177-R185del patient and an unaffected family member. To evaluate the therapeutic efficacy of MANF, we generated iPSC lines with inducible MANF overexpression (OE) and differentiated them into kidney organoids. TALs were isolated from these organoids via Magnetic bead conjugation system for analysis by WB/RNA-seq. ER calcium dynamics were assessed using confocal microscopy and a novel secreted ER calcium-monitoring protein (SERCaMP). Additionally, protein interaction domains were mapped by deletion mutagenesis and Co-IP. Finally, secreted inflammatory cytokines were quantified using a newly invented, ultrasensitive plasmon-enhanced fluorescence-linked immunosorbent multiplex assay (p-FLISA).

Results

The mutant organoids recapitulate key disease features, including mutant UMOD-triggered ER stress activation and defective p-AMPK/FOXO3-mediated autophagy. Furthermore, mutant TALs isolated from the ADTKD organoids exhibit marked ER Ca2+ leakage driven by the selective upregulation of ER Ca2+ release channel IP3R1, which in turn induces tribbles homolog 3 (TRIB3). Mechanistically, TRIB3 directly interacts with the activation loop of AMPKα, suppressing its phosphorylation at T183/T172 and thereby impairing autophagy. MANF OE directly binds to IP3R1, mitigating ER Ca2+ efflux and subsequent TRIB3 induction. The subsequent recovery of AMPK activity enhances autophagic clearance of mutant UMOD. Consequently, MANF OE attenuates ER stress-associated inflammation and TAL apoptosis, ultimately promoting mutant TAL recovery.

Conclusion

MANF restores autophagy by blunting aberrant ER Ca2+ efflux in ADTKD organoids. Thus, MANF represents a potent therapy to correct proteostasis in ADTKD and potentially other protein misfolding diseases.

Acknowledgment

C.J.G is supported by American Society of Nephrology (ASN) Kidney Cure Ben J. Lipps Research Fellowship Program. Y.L.F is supported by the American Heart Association Postdoctoral Fellowship. Y.M.C. is supported by NIH grants R01 2DK105056-6, R56DK138158A1, R21DK131557A1, R03DK106451 and K08DK089015, VA Merit I01BX006401A1.

Funding

  • NIDDK Support