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Influence of Hydrolyzed Polyacrylamide Hydrogel Stiffness on Podocyte Morphology, Phenotype, and Mechanical Properties.

ACS applied materials & interfaces (2019-08-20)
Maya Abdallah, Marta Martin, Mario R El Tahchi, Sebastien Balme, Wissam H Faour, Béla Varga, Thierry Cloitre, Orsolya Páll, Frédéric J G Cuisinier, Csilla Gergely, Maria J Bassil, Mikhael Bechelany
ABSTRACT

Chronic kidney disease is characterized by a gradual decline in renal function that progresses toward end-stage renal disease. Podocytes are highly specialized glomerular epithelial cells which form with the glomerular basement membrane (GBM) and capillary endothelium the glomerular filtration barrier. GBM is an extracellular matrix (ECM) that acts as a mechanical support and provides biophysical signals that control normal podocytes behavior in the process of glomerular filtration. Thus, the ECM stiffness represents an essential characteristic that controls podocyte function. Hydrolyzed Polyacrylamide (PAAm) hydrogels are smart polyelectrolyte materials. Their biophysical properties can be tuned as desired to mimic the natural ECM. Therefore, these hydrogels are investigated as new ECM-like constructs to engineer a podocyte-like basement membrane that forms with cultured human podocytes a functional glomerular-like filtration barrier. Such ECM-like PAAm hydrogel construct will provide unique opportunity to reveal podocyte cell biological responses in an in vivo-like setting by controlling the physical properties of the PAAm membranes. In this work, Hydrolyzed PAAm scaffolds having different stiffness ranging between 0.6-44 kPa are prepared. The correlation between the hydrogel structural and mechanical properties and Podocyte morphology, elasticity, cytoskeleton reorganization, and podocin expression is evaluated. Results show that hydrolyzed PAAm hydrogels promote good cell adhesion and growth and are suitable materials for the development of future 3D smart scaffolds. In addition, the hydrogel properties can be easily modulated over a wide physiological range by controlling the cross-linker concentration. Finally, tuning the hydrogel properties is an effective strategy to control the cells function. This work addressed the complexity of podocytes behavior which will further enhance our knowledge to develop a kidney-on-chip model much needed in kidney function studies in both healthy and diseased states.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
RPMI-1640 Medium, With L-glutamine and sodium bicarbonate, liquid, sterile-filtered, suitable for cell culture
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Penicillin-Streptomycin, Solution stabilized, with 10,000 units penicillin and 10 mg streptomycin/mL, 0.1 μm filtered, BioReagent, suitable for cell culture
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DAPI, for nucleic acid staining
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N-Hydroxysuccinimide, 98%
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Thiazolyl Blue Tetrazolium Bromide, powder, BioReagent, suitable for cell culture, suitable for insect cell culture, ≥97.5% (HPLC)
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N-(3-Dimethylaminopropyl)-N′-ethylcarbodiimide hydrochloride, crystalline
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Ammonium persulfate, ACS reagent, ≥98.0%
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Acrylamide, suitable for electrophoresis, ≥99%
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N,N′-Methylenebisacrylamide, powder, for molecular biology, suitable for electrophoresis, ≥99.5%
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N,N,N′,N′-Tetramethylethylenediamine, BioReagent, for molecular biology, ≥99% (GC)
Sigma-Aldrich
Anti-Podocin antibody produced in rabbit, affinity isolated antibody, buffered aqueous solution
Sigma-Aldrich
Poly(trimethylene carbonate), viscosity 1.75 dL/g 

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