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Abstract: FR-PO820

Akt Downregulation Induces Tubular Apoptosis via FoxO-1-Induced BIM Activation in Proteinuric States

Session Information

Category: Glomerular Diseases

  • 1202 Glomerular Diseases: Immunology and Inflammation

Authors

  • Erkan, Elif, Children's Hospital of Cincinnati, CIncinnati, Ohio, United States
  • Schuh, Meredith Posner, Cincinnati Children's Hospital Medical Center, Cincinnati, Ohio, United States
Background

Proteinuria induces tubular apoptosis preceding tubular atrophy but the underlying molecular mechanism remains undetermined. We demonstrated that cell survival protein, protein kinase B(Akt) is downregulated in proximal tubule epithelial cells in response to albumin overload. We hypothesize that inhibition of Akt expression decreases phosporylation and activation of its downstream targets Forkhead box(FoxO) transcriptional factors leading to mitochondrial apoptosis in proteinuric states.

Methods

In-vitro albumin overload: Human kidney proximal tubule epithelial cells(HKC-8) were incubated with 10mg/ml endotoxin free human albumin for 6, 16 and 24 hours.
Chromatin immunoprecipitation(CHIP) assay was used to probe protein-DNA interactions.
In-vivo albumin overload: Wild type and Akt1/2 lox/lox SGLT2 cre+ mice underwent daily intraperitoneal albumin injections(10mg/g) for 6 weeks.
Human kidney biopsies with minimal change disease and focal segmental glomerulosclerosis (FSGS) were investigated for Akt activation.

Results

Expression of phosphorylated Akt-Ser 473 and Akt-Thr 308 was downregulated in association with increased caspase-3 activity and BIM expression in HKC-8 cells with albumin overload. Treatment of HKC-8 cells with pan Akt inhibitor MK-2206 and constitutively active(CA-Akt) Akt resulted in down regulation and upregulation of apoptosis respectively with albumin overload.
In-vivo albumin overload decreased active p-Akt expression in association with tubular apoptosis. Akt1/2 lox/lox SGLT2 cre+ mice displayed increased BIM/Bax expression in mitochondrial isolates in association with tubular apoptosis in response to albumin overload indicating a close causal link between inhibition of Akt and mitochondrial apoptosis in proximal tubule epithelial cells. Furthermore, patient kidney biopsies (n=5) with FSGS displayed decreased proximal tubule PSer473 Akt expression.
Albumin overload caused decreased Akt phosphorylation of FoxO-1 and 3 and nuclear translocation of FoxO-1. CHIP assay revaled transcriptional activation of BIM by Foxo-1.

Conclusion

We concluded that Akt and its downstream targets play an important role in mediating mitochondrial apoptosis leading to tubular injury in glomerular diseases. We propose that discovery of novel treatment options targeted for activation of Akt can ameliorate tubulointerstitial injury and progression in proteinuric states.