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Nephrol Dial Transplant. 2019 May 23. pii: gfz094. doi: 10.1093/ndt/gfz094. [Epub ahead of print]

Towards an in vitro model of glomerular barrier unit with an innovative bioassembly method.

Flegeau K1,2, Rubin S1,2,3, Mucha S1,2,3, Bur P1,2, Préterre J1,2, Siadous R1,2, L'Azou B1,2, Fricain JC1,2,4, Combe C1,2,3, Devillard R1,2,4, Kalisky J1,2, Rigothier C1,2,3.

Author information

1
Tissue Bioengineering, Université de Bordeaux, Bordeaux, France.
2
Tissue Bioengineering, INSERM, Bordeaux, France.
3
Service de Néphrologie Transplantation, Dialyse et Aphérèse, Centre Hospitalier Universitaire de Bordeaux, Bordeaux, France.
4
Service d'odontologie et de Santé Buccale, Centre Hospitalier Universitaire de Bordeaux, Bordeaux, France.

Abstract

BACKGROUND:

The development of an artificial glomerular unit may be pivotal for renal pathophysiology studies at a multicellular scale. Using a tissue engineering approach, we aimed to reproduce in part the specific glomerular barrier architecture by manufacturing a glomerular microfibre (Mf).

METHODS:

Immortalized human glomerular cell lines of endothelial cells (GEnCs) and podocytes were used. Cells and a three-dimensional (3D) matrix were characterized by immunofluorescence with confocal analysis, Western blot and polymerase chain reaction. Optical and electron microscopy were used to study Mf and cell shapes. We also analysed cell viability and cell metabolism within the 3D construct at 14 days.

RESULTS:

Using the Mf manufacturing method, we repeatedly obtained a cellularized Mf sorting human glomerular cells in 3D. Around a central structure made of collagen I, we obtained an internal layer composed of GEnC, a newly formed glomerular basement membrane rich in α5 collagen IV and an external layer of podocytes. The cell concentration, optimal seeding time and role of physical stresses were modulated to obtain the Mf. Cell viability and expression of specific proteins (nephrin, synaptopodin, vascular endothelial growth factor receptor 2 (VEGFR2) and von Willebrandt factor (vWF)) were maintained for 19 days in the Mf system. Mf ultrastructure, observed with EM, had similarities with the human glomerular barrier.

CONCLUSION:

In summary, with our 3D bio-engineered glomerular fibre, GEnC and podocytes produced a glomerular basement membrane. In the future, this glomerular Mf will allow us to study cell interactions in a 3D system and increase our knowledge of glomerular pathophysiology.

KEYWORDS:

bioassembly; glomerular barrier; glomerular endothelial cells; podocytes; tissue engineering

PMID:
31121032
DOI:
10.1093/ndt/gfz094

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