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Abstract: SA-PO370

A Novel Mouse Model Embodying Minimal-Change Nephrotic Syndrome Through Crumbs 2-Mediated Signaling to the Cytoskeleton in Podocytes

Session Information

Category: Glomerular Diseases

  • 1202 Glomerular Diseases: Immunology and Inflammation

Authors

  • Hada, Ichiro, Kyorin University, Mitaka, ToKyo, Japan
  • Mikami, Naoaki, Kyorin University, Mitaka, ToKyo, Japan
  • Mori, Satoka, Denka Co., Ltd., Tokyo, Japan
  • Hirayama, Yoshiaki, Denka-seiken co., ltd., Gosen-shi, Japan
  • Shimizu, Akira, Nippion Medical School, Tokyo, Japan
  • Yan, Kunimasa, Kyorin University School of Medicine, Mitaka, ToKyo, Japan
Background

The cellular events in podocyte injury causing minimal-change nephrotic syndrome (MCNS) remain largely obscure. Recent reports implicated genetic mutation of crumbs homolog 2 (CRB2), a type-1 transmembrane protein, as a cause of congenital nephrotic syndrome. However, an involvement of CRB2 dysfunction in the selective permeability barrier of mature glomerulus has not been demonstrated. The aim of the present study is to determine whether MCNS is caused by alteration of CRB2-mediated signaling in podocytes.

Methods

A partial recombinant protein including the extracellular part of mouse CRB2 was produced in an Escherichia coli system. An Enzyme-Linked Immunosorbent Assay was established to measure anti-CRB2 antibody in the serum. C3H/HeNmice and A/Jmice (complement C5 deficient mice) were injected with CRB2 recombinant protein. Urinary protein was analyzed, and kidney tissue was subjected to immunofluorescence microscopy, light microscopy, electron microscopy and immuno-electron microscopy of actin. Isolated glomeruli were subjected to Western blot analysis of phosphorylated moesin.

Results

Both C3H/HeNmice and A/J mice developed massive proteinuria at 4-6 weeks after immunization with CRB recombinant protein, which was associated with elevation of anti-CRB2 antibody. Light microscopy revealed minimal glomerular lesions even at a heavily proteinuric stage. Electron microscopy revealed prominent foot process effacement where broad dense areas were seen. Immuno-electron microscopy showed that these areas were positively immunostained for actin. Immunofluorescence microscopy revealed positive IgG staining in podocyte foot processes, but not complement C3. Finally, Western blot analysis showed phosphorylation of moesin in glomerular samples at the proteinuric stage.

Conclusion

The present study reveals a novel pathway involved in the pathophysiology of MCNS through an interaction between CRB2-mediated signaling and the actin cytoskeleton network. CRB2 may be an important scaffold molecule that mediates outside-in signaling in podocytes, thereby leading to actin cytoskeleton reorganization and resulting in MCNS.