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Isolation of Primary Rat Liver Fibroblasts
  1. Emma A. Kruglov,
  2. Dhanpat Jain,
  3. Jonathan A. Dranoff
  1. From the Department of Medicine and Yale Liver Center (E.A.K., J.A.D.), Yale University School of Medicine, New Haven, Conn.
  2. Department of Pathology (D.J.), Yale University School of Medicine, New Haven, Conn.
  1. Address correspondence to: Jonathan A. Dranoff, MD, Gastroenterology Research Lab, VAMC Connecticut, 950 Campbell Avenue, Bldg. 27, West Haven, CT 06520. Email: jonathan.dranoff{at}yale.edu
  2. This work was supported by Grant No. DK02379 from the National Institutes of Health to J.A.D., a Basic Research Award from the American Association for the Study of Liver Diseases to J.A.D., and Grant No. DK34989 from the National Institutes of Health to the Yale Liver Center.

Abstract

Introduction One of the major advances in liver research in the past decade was the ability to isolate distinct liver cell populations. Although there are established methods of isolating hepatocytes, cholangiocytes, and stellate cells, before this study no technique for liver fibroblast isolation had been devised. Consequently, we developed a technique to isolate primary rat liver fibroblasts.

Methods Fibroblasts were isolated from a freshly perfused rat liver with a modification of the procedure for isolation of rat cholangiocytes. Cell markers were assessed with the use of confocal immunofluorescence. Cell morphology was assessed with transmission electron microscopy. Expression of procollagen-1 was assessed by reverse transcription polymerase chain reaction.

Results The appearance of cells with fibroblast morphology was first noted at 48 hours, and almost all cells in culture had fibroblast morphology at 96 hours. Putative fibroblasts stained for vimentin, but not for smooth muscle actin, von Willebrand factor, or cytokeratins. Cell morphology was consistent with that of fibroblasts and showed no features of epithelial, endothelial, or smooth muscle cells. Liver fibroblasts expressed procollagen-1 mRNA.

Conclusion Primary isolated rat fibroblasts can be produced from a freshly perfused rat liver with a modification of standard cell culture methods. The role of fibroblasts in liver physiology can now be studied directly.

Key Words
  • cell isolation
  • fibroblast
  • liver
  • nonparenchymal cell

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