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Kidney Week

Abstract: FR-OR072

Rho Guanine Nucleotide Exchange Factor β-PIX Is Required for the Maintenance of Podocyte Architecture and Glomerular Function in Adult Mice

Session Information

Category: Genetic Diseases of the Kidney

  • 1002 Genetic Diseases of the Kidney: Non-Cystic

Authors

  • Matsuda, Jun, McGill University Health Centre, Montreal, Quebec, Canada
  • Maier, Mirela, McGill University Health Centre, Montreal, Quebec, Canada
  • Aoudjit, Lamine, McGill University Health Centre, Montreal, Quebec, Canada
  • Takano, Tomoko, McGill University Health Centre, Montreal, Quebec, Canada
Background

Activation of Rac1 (a small GTPase) in podocytes has been implicated in the development of proteinuria and focal segmental glomerulosclerosis (FSGS). Previously, we identified Arhgef7 (β-PIX) as a predominant GEF (guanine nucleotide exchange factor) that interacts with Rac1 in human podocytes (HP) using a proximity-based ligation assay, BioID. Furthermore, we confirmed that β-PIX is expressed in podocytes both in vitro and in vivo. However, its functional role remains unknown.

Methods

Cultured mouse podocytes (MP) with β-PIX knockdown (KD) and their controls were established using shRNA. Cell size was quantified in fixed cells stained with phalloidin. Cell migration and adhesive property were studied by wound healing assay and attachment assay, respectively. Cell proliferation was studied by MTT assay. Podocyte specific β-PIX deficient (KO) mice were generated by crossing β-PIX floxed mice with NPHS2-Cre mice. Urine albumin to creatinine ratio (ACR) and renal histology were studied up to 13 weeks of age. Data are provided as the mean ± SE.

Results

Compared to control MP, β-PIX KD MP were significantly smaller. Cell motility and adhesive property were modestly decrease in β-PIX KD MP. Proliferation rate was not different. Podocyte specific β-PIX KO mice presented no obvious renal phenotype up to 2-3 weeks of age, but developed progressive proteinuria starting at around 7-8 weeks of age (8 wk: KO: 117±44 x102 vs Control: 49±15 μg/mg, n=7-11, p<0.01; 13 wk: 255±128 x102 vs 238±90 μg/mg, n=4-7, p<0.05). In addition, by PAS staining, 13-week-old KO mice showed global/segmental glomerulosclerosis, parietal cells proliferation, and tubular casts in the kidney.

Conclusion

Our findings demonstrate the previously unrecognized importance of β-PIX in maintaining podocyte function. To the best of our knowledge, this is the first evidence that a Rho GTPase GEF plays a critical role in podocytes.