Geltrex‐Enhanced Two‐Dimensional Culture as a Viable Alternative to Primary Rat Hepatocyte Sandwich Models
Eda Beldek, Matilda Holtz, Adil Denizli, O. B. Usta
- Authors
- Eda Beldek, Matilda Holtz, Adil Denizli, O. B. Usta
- Published
- Comprehensive Physiology 2025
- DOI
- 10.1002/cph4.70082
- Citations
- 0
Abstract
Primary hepatocytes rapidly lose viability and function in conventional two‐dimensional (2D) cultures due to the absence of a physiologically relevant extracellular matrix (ECM). The collagen sandwich method improves polarization and function but creates a diffusion barrier that limits nutrient and signal exchange. This study investigates whether daily supplementation of a diluted, non‐gelling Geltrex layer can sustain hepatocyte function and viability in 2D culture, offering a practical alternative to the sandwich method. Primary rat hepatocytes were cultured for 15 days under four conditions: monolayer (ML), monolayer with Geltrex (ML + GT), sandwich (SW), and sandwich with Geltrex (SW + GT). Cell morphology, confluency, viability (CCK‐8, live/dead staining), and functionality (urea synthesis, albumin production, CYP3A4 activity) were assessed. The ML group showed significant declines in confluency, viability, and functional markers over time. Geltrex supplementation preserved confluency (~97% at day 15), improved viability, and maintained higher albumin production and CYP3A4 activity compared to ML. Functional outputs in ML + GT were comparable to SW and SW + GT groups, without the diffusion limitations of the sandwich top gel. Daily supplementation with low‐dose Geltrex creates a biochemically enriched, diffusion‐permissive microenvironment that supports long‐term viability and function of primary rat hepatocytes in 2D culture. This method represents a simple and effective alternative to traditional sandwich cultures for liver cell studies and drug testing applications.
Cite this article
Eda Beldek, Matilda Holtz, Adil Denizli, O. B. Usta (2025). Geltrex‐Enhanced Two‐Dimensional Culture as a Viable Alternative to Primary Rat Hepatocyte Sandwich Models. Comprehensive Physiology. https://doi.org/10.1002/cph4.70082
BibTeX
@article{eda2025,
title = {Geltrex‐Enhanced Two‐Dimensional Culture as a Viable Alternative to Primary Rat Hepatocyte Sandwich Models},
author = {Eda Beldek and Matilda Holtz and Adil Denizli and O. B. Usta},
journal = {Comprehensive Physiology},
doi = {10.1002/cph4.70082},
year = {2025}
}