Abstract: TH-PO424
Pressure-Dependent Molecular Transport in Native and Enzymatically Crosslinked Glomerular Basement Membrane
Session Information
- Glomerular Diseases: Inflammation and Fibrosis
November 03, 2022 | Location: Exhibit Hall, Orange County Convention Center‚ West Building
Abstract Time: 10:00 AM - 12:00 PM
Category: Glomerular Diseases
- 1301 Glomerular Diseases: Fibrosis and Extracellular Matrix
Authors
- Wang, Dan, The Ohio State University Wexner Medical Center, Columbus, Ohio, United States
- Ferrell, Nicholas J., The Ohio State University Wexner Medical Center, Columbus, Ohio, United States
Background
Glomerular basement membrane (GBM) is a thin layer of extracellular matrix (ECM) that supports podocytes and endothelial cells and regulates their behavior and function. The role of the GBM in determining the permselectivity of the glomerular filter in health and disease is not fully understood. The GBM has been proposed to act as a compressible membrane with increased selectivity under applied pressure. Multiple kidney diseases are characterized by increased enzymatic crosslinking of the ECM. We hypothesized that enzymatic crosslinking would increase GBM permeability under pressure by increasing GBM stiffness and reducing pressure dependent compression.
Methods
Glomeruli were isolated from porcine kidneys by sieving. Stiffness of native and microbial transglutaminase (mTG) crosslinked decellularized glomeruli were measured by a customized compression assay. GBM membranes were made by pressure compacting decellularized glomeruli on a supporting membrane in a stirred cell. FITC-Ficoll sieving coefficients were measured at high and low pressure on native and crosslinked GBM. Diffusional permeability was determined by measuring passive transport of Ficoll through the GBM in the absence of applied pressure.
Results
Stiffness of decellularized glomeruli was significantly increased (4.1-fold) after crosslinking with mTG (100 μg/mL). mTG did not affect FITC-Ficoll diffusional permeability of GBM, suggesting minimal alteration of pore structure in the absence of applied pressure. The sieving coefficient curves of native GBM show a reduction in molecular cut-off with increasing pressure, indicative of membrane compression effects. Crosslinked membranes did not show significant compression effects on sieving coefficient.
Conclusion
We developed a GBM in vitro model to investigate the molecular transport. We use the native and crosslinked GBM as healthy and pathological conditions, respectively. mTG had minimal effect on molecular transport during diffusive transport. Native GBM behaved as a compressive filter, restricting large molecule transport at high pressure. This effect was mitigated in enzymatically crosslinked GBM due to the decreased degree of compressibility suggesting that disease mediated enzymatic crosslinking may contribute to glomerular filtration defects in chronic kidney disease.
Funding
- Other U.S. Government Support