Isolation, long-term culture, and characterization of rat pancreatic fibroblastoid/stellate cells

Pancreas. 2001 Jul;23(1):49-54. doi: 10.1097/00006676-200107000-00007.

Abstract

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.

MeSH terms

  • Actins / analysis
  • Animals
  • Azo Compounds
  • Biomarkers
  • Blotting, Western
  • Cell Culture Techniques / methods*
  • Cell Differentiation
  • Cell Line
  • Cell Separation
  • Ceruletide / toxicity
  • Coloring Agents
  • Contact Inhibition
  • Culture Media
  • Desmin / analysis
  • Fibroblasts / cytology*
  • Fluorescent Antibody Technique, Indirect
  • Lipids / analysis
  • Male
  • Muscle, Smooth / chemistry
  • Muscle, Smooth / ultrastructure
  • Pancreas / cytology*
  • Pancreas / drug effects
  • Pancreatitis / chemically induced
  • Pancreatitis / metabolism
  • Pancreatitis / pathology
  • Rats
  • Rats, Wistar
  • Staining and Labeling

Substances

  • Actins
  • Azo Compounds
  • Biomarkers
  • Coloring Agents
  • Culture Media
  • Desmin
  • Lipids
  • Ceruletide
  • oil red O