Primary rat hepatocyte culture on 3D nanofibrous polymer scaffolds for toxicology and pharmaceutical research.
Primary rat hepatocytes are a widely used experimental model to estimate drug metabolism and toxicity. In currently used two-dimensional (2D) cell culture systems, typical problems like morphological changes and the loss of liver cell-specific functions occur. We hypothesize that the use of polymer scaffolds could overcome these problems and support the establishment of three-dimensional (3D) culture systems in pharmaceutical research. Isolated primary rat hepatocytes were cultured on collagen-coated nanofibrous scaffolds for 7 days. Cell loading efficiency was quantified via DNA content measurement. Cell viability and presence of liver-cell-specific functions (albumin secretion, glycogen storage capacity) were evaluated. The activity of liver-specific factors was analyzed by immunofluorescent staining. RNA was isolated to establish quantitative real-time PCR. Our results indicate that primary rat hepatocytes cultured on nanofibrous scaffolds revealed high viability and well-preserved glycogen storage. Albumin secretion was existent during the entire culture period. Hepatocytes remain HNF-4 positive, indicating highly preserved cell differentiation. Aggregated hepatocytes re-established positive signaling for Connexin 32, a marker for differentiated hepatocyte interaction. ZO-1-positive hepatocytes were detected indicating formation of tight junctions. Expression of cytochrome isoenzymes was inducible. Altogether the data suggest that nanofibrous scaffolds provide a good in vitro microenvironment for neo tissue regeneration of primary rat hepatocytes.
Bierwolf
Jeanette
J
Department of Hepatobiliary and Transplant Surgery, University Medical Center Hamburg-Eppendorf, Martinistrasse 52, 20246 Hamburg, Germany.
Lutgehetmann
Marc
M
Feng
Kai
K
Erbes
Johannes
J
Deichmann
Steffen
S
Toronyi
Eva
E
Stieglitz
Christina
C
Nashan
Bjoern
B
Ma
Peter X
PX
Pollok
Joerg M
JM
D000818:Animals; D002470:Cell Survival; D002478:Cells, Cultured; D022781:Hepatocytes / Q000502:physiology*; D019164:Liver, Artificial; D057139:Nanofibers; D009924:Organ Culture Techniques; D010600:Pharmacology / Q000379:methods; D011108:Polymers; D051381:Rats; D023822:Tissue Engineering / Q000379:methods; D054457:Tissue Scaffolds; D014116:Toxicology / Q000379:methods
D011108:Polymers