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A Cell Culture System for the Structure and Hydrogel Properties of Basement Membranes: Application to Capillary Walls

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Abstract

In specialized capillary beds such as the kidney glomerulus, the sheet-like structure of the basement membrane (BM) in conjunction with opposing monolayers of endothelium and epithelium form the functioning filtration unit of the kidney. Using a novel cross-linking method on a collagen substrate, we have created a hydrogel scaffold to substitute for the BM. A simple casting method was used to create thin films of the hydrogel scaffold (1–5 μm), that were suitable for long-term static culture, and supported cell attachment and long term cell viability similar to a standard type I collagen substrate. Bulk diffusion and protein permeability of the hydrogel scaffold were evaluated, in addition to its use in a perfusion chamber where it withstood hydraulic pressures typical for glomerular capillaries. This system thus provided a suitable cell substrate for the co-culture of renal epithelial podocytes and endothelial cells in a device that replicates the geometry of the in vivo juxtaposition of the two cell types in relation to their BM.

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Abbreviations

BM:

Basement membrane

BSA:

Bovine serum albumin

HRP:

Horseradish peroxidase

T-gelatin:

Tyramine-substituted gelatin

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Acknowledgments

We thank Dr. Michael Madaio for providing the endothelial cell line, Zhenzhen Wu for technical assistance, and Sydney Calabro for assistance with the images. We also acknowledge the helpful input from Drs. Jeffrey Simske, John Sedor, Tyler Miller, and William Fissell for discussions and review of the manuscript. This study was supported by NIH grant DK077668 and MetroHealth Medical Center institutional funds.

Conflict of interest

The authors (AC and AD) have licensed the tyramine substitution and cross-linking technology for commercialization to Lifecore Biomedical, Inc., Chaska, MN and have received benefits for personal or professional use indirectly related to the subject matter of this manuscript. Benefits have been used for research funding at The Cleveland Clinic, a nonprofit organization, with which these authors are associated.

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Author notes

  1. Ryan P. Doan

    Present address: Santa Clara Valley Medical Center, Stanford University School of Medicine, San Jose, CA, 95128, USA

  2. Aniq Darr

    Present address: Rutgers University, Piscataway, NJ, 08854, USA

Authors and Affiliations

  1. Department of Medicine, Rammelkamp Center for Education and Research, MetroHealth Medical Center, Case Western Reserve University School of Medicine, Rammelkamp Building, Room R429, 2500 MetroHealth Drive, Cleveland, OH, 44109, USA

    Leslie A. Bruggeman

  2. Cleveland Clinic Lerner College of Medicine, Lerner Research Institute, The Cleveland Clinic, Cleveland, OH, 44195, USA

    Ryan P. Doan & Anthony Calabro

  3. Department of Biomedical Engineering, Lerner Research Institute, The Cleveland Clinic, Cleveland, OH, 44195, USA

    Jacqueline Loftis, Aniq Darr & Anthony Calabro

Authors
  1. Leslie A. Bruggeman
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  2. Ryan P. Doan
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  3. Jacqueline Loftis
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  4. Aniq Darr
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  5. Anthony Calabro
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Corresponding author

Correspondence to Leslie A. Bruggeman.

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Associate Editor Muhammad Zaman oversaw the review of this article.

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Bruggeman, L.A., Doan, R.P., Loftis, J. et al. A Cell Culture System for the Structure and Hydrogel Properties of Basement Membranes: Application to Capillary Walls. Cel. Mol. Bioeng. 5, 194–204 (2012). https://doi.org/10.1007/s12195-012-0221-3

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  • DOI: https://doi.org/10.1007/s12195-012-0221-3

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