Isolation, long-term culture, and characterization of rat pancreatic fibroblastoid/stellate cells.
Investigation of pancreatic interstitial fibroblasts has proven difficult in situ. We have established a method for the isolation of pancreatic fibroblastoid/stellate cells by outgrowth from pancreatic tissue explanted into culture dishes. This technique gives a high yield of viable cells from small tissue samples. Outgrown fibroblastoid cells were established as a primary cell line and characterized during long-term culture. We investigated the development of stellate cell markers, i.e. fat storage, expression of desmin, and alpha-smooth muscle actin (alphaSMA), over weeks in culture. AlphaSMA, investigated by indirect immunofluorescence staining and Western blot analysis, revealed a constant rise in expression during routine culture. After 13 passages. approximately 100% of cells were positive for alphaSMA expression, indicating a myofibroblast type of differentiation in vitro.
Kruse
M L
ML
Laboratory for Molecular Gastroenterology and Hepatology, 1st Department of Medicine, University of Kiel, Germany. mlkruse@1med.uni-kiel.de
Hildebrand
P B
PB
Timke
C
C
Fölsch
U R
UR
Schäfer
H
H
Schmidt
W E
WE
D000199:Actins / Q000032:analysis; D000818:Animals; D001391:Azo Compounds; D015415:Biomarkers; D015153:Blotting, Western; D018929:Cell Culture Techniques / Q000379:methods*; D002454:Cell Differentiation; D002460:Cell Line; D002469:Cell Separation; D002108:Ceruletide / Q000633:toxicity; D004396:Coloring Agents; D003260:Contact Inhibition; D003470:Culture Media; D003893:Desmin / Q000032:analysis; D005347:Fibroblasts / Q000166:cytology*; D019084:Fluorescent Antibody Technique, Indirect; D008055:Lipids / Q000032:analysis; D008297:Male; D009130:Muscle, Smooth / Q000737:chemistry / Q000648:ultrastructure; D010179:Pancreas / Q000166:cytology* / Q000187:drug effects; D010195:Pancreatitis / Q000139:chemically induced / Q000378:metabolism / Q000473:pathology; D051381:Rats; D017208:Rats, Wistar; D013194:Staining and Labeling
D000199:Actins; D001391:Azo Compounds; D015415:Biomarkers; D004396:Coloring Agents; D003470:Culture Media; D003893:Desmin; D008055:Lipids; D002108:Ceruletide; C011049:oil red O