Fibrosis pp 165-191 | Cite as

Isolation and Culture of Primary Murine Hepatic Stellate Cells

Part of the Methods in Molecular Biology book series (MIMB, volume 1627)


Hepatic stellate cells (HSCs) are found in the perisinusoidal space of the liver (i.e., the space of Dissé). They represent 5–8% of the total number of liver cells. In normal liver, these cells have a quiescent phenotype and are characterized by numerous fat vacuoles that store vitamin A in a form of retinyl ester. In injured liver, these cells transdifferentiate into a myofibroblast phenotype, become highly proliferative and are responsible for excess collagen synthesis and deposition during fibrosis. Due to their exceptional pathophysiological relevance, several isolation and purification protocols of primary HSCs have been established that provide the basis for studying HSC biology in vitro. We here describe a method for high-purity isolation of HSCs from mice. This protocol includes the enzymatic digestion of the liver tissue by pronase and collagenase, cellular enrichment by centrifugation of the crude cell suspension through a Nycodenz density gradient, and a final (optional) flow cytometric enrichment that allows generating ultrapure HSC fractions.

Key words

Hepatic stellate cells Myofibroblasts Nycodenz FACS Density gradient Desmin Phalloidin 



This work was supported by grants from the German Research Foundation (SFB/TRR 57 projects P09, P13, and Q3).


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© Springer Science+Business Media LLC 2017

Authors and Affiliations

  1. 1.Institute of Molecular Pathobiochemistry, Experimental Gene Therapy and Clinical ChemistryRWTH University Hospital AachenAachenGermany
  2. 2.Department of Internal Medicine IIIRWTH University Hospital AachenAachenGermany

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