Genetic and Molecular Characterization of the Immortalized Murine Hepatic Stellate Cell Line GRX

Sarah K Schröder; Herdit M Schüler; Kamilla V Petersen; Cinzia Tesauro; Birgitta R Knudsen; Finn S Pedersen; Frederike Krus; Eva M Buhl; Elke Roeb; Martin Roderfeld; Radovan Borojevic; Jamie L Almeida; Ralf Weiskirchen

doi:10.3390/cells11091504

Genetic and Molecular Characterization of the Immortalized Murine Hepatic Stellate Cell Line GRX

Cells. 2022 Apr 30;11(9):1504. doi: 10.3390/cells11091504.

Authors

Affiliations

¹ Institute of Molecular Pathobiochemistry, Experimental Gene Therapy and Clinical Chemistry (IFMPEGKC), RWTH University Hospital Aachen, D-52074 Aachen, Germany.
² Institute of Human Genetics, University Hospital of RWTH Aachen University, D-52074 Aachen, Germany.
³ Department of Molecular Biology and Genetics, Aarhus University, DK-8000 Aarhus, Denmark.
⁴ Department of Clinical Medicine, Aarhus University, DK-8000 Aarhus, Denmark.
⁵ VPCIR Biosciences ApS., DK-8000 Aarhus, Denmark.
⁶ Electron Microscopy Facility, Institute of Pathology, RWTH Aachen University Hospital, D-52074 Aachen, Germany.
⁷ Department of Gastroenterology, Justus Liebig University, D-35392 Giessen, Germany.
⁸ Institute of Biomedical Sciences, Federal University of Rio de Janeiro, Rio de Janeiro 21941-590, Brazil.
⁹ National Institute of Standards and Technology, Biosystems and Biomaterials Division, Gaithersburg, MD 20899, USA.

Abstract

The murine cell line GRX has been introduced as an experimental tool to study aspects of hepatic stellate cell biology. It was established from livers of C3H/HeN mice that were infected with cercariae of Schistosoma mansoni. Although these cells display a myofibroblast phenotype, they can accumulate intracellular lipids and acquire a fat-storing lipocyte phenotype when treated with retinol, insulin, and indomethacin. We have performed genetic characterization of GRX and established a multi-loci short tandem repeat (STR) signature for this cell line that includes 18 mouse STR markers. Karyotyping further revealed that this cell line has a complex genotype with various chromosomal aberrations. Transmission electron microscopy revealed that GRX cells produce large quantities of viral particles belonging to the gammaretroviral genus of the Retroviridae family as assessed by next generation mRNA sequencing and Western blot analysis. Rolling-circle-enhanced-enzyme-activity detection (REEAD) revealed the absence of retroviral integrase activity in cell culture supernatants, most likely as a result of tetherin-mediated trapping of viral particles at the cell surface. Furthermore, staining against schistosome gut-associated circulating anodic antigens and cercarial O- and GSL-glycans showed that the cell line lacks S. mansoni-specific glycostructures. Our findings will now help to fulfill the recommendations for cellular authentications required by many granting agencies and scientific journals when working with GRX cells. Moreover, the definition of a characteristic STR profile will increase the value of GRX cells in research and provides an important benchmark to identify intra-laboratory cell line heterogeneity, discriminate between different mouse cell lines, and to avoid misinterpretation of experimental findings by usage of misidentified or cross-contaminated cells.

Keywords: 3R principle; STR profiling; authentication; cell line; cross-contamination; genotyping; hepatic stellate cells; liver; myofibroblasts; retinol.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Animals
Hepatic Stellate Cells* / metabolism
Kupffer Cells* / metabolism
Liver / metabolism
Mice
Mice, Inbred C3H
Vitamin A / metabolism

Substances

Vitamin A

Grants and funding

R.W. and M.R. are financially supported by grants from the German Research Foundation (projects WE2554/13-1, W2554/15-1, WE 2554/17-1, and RO 3714/4-1). The funder had no role in the design of this article or in the decision to publish it.