Comparison of the Opn-CreER and Ck19-CreER Drivers in Bile Ducts of Normal and Injured Mouse Livers

Lesaffer Lesaffer,Van Grunsven Van Grunsven,Leclercq Leclercq,Verboven Verboven,Van Huffel Van Huffel,Moya Moya,Halder Halder,Lemaigre Lemaigre

doi:10.3390/cells8040380

Abstract

Inducible cyclization recombinase (Cre) transgenic mouse strains are powerful tools for cell lineage tracing and tissue-specific knockout experiments. However, low efficiency or leaky expression can be important pitfalls. Here, we compared the efficiency and specificity of two commonly used cholangiocyte-specific Cre drivers, the Opn-iCreERT2 and Ck19-CreERT drivers, using a tdTomato reporter strain. We found that Opn-iCreERT2 triggered recombination of the tdTomato reporter in 99.9% of all cholangiocytes while Ck19-CreERT only had 32% recombination efficiency after tamoxifen injection. In the absence of tamoxifen, recombination was also induced in 2% of cholangiocytes for the Opn-iCreERT2 driver and in 13% for the Ck19-CreERT driver. For both drivers, Cre recombination was highly specific for cholangiocytes since recombination was rare in other liver cell types. Toxic liver injury ectopically activated Opn-iCreERT2 but not Ck19-CreERT expression in hepatocytes. However, ectopic recombination in hepatocytes could be avoided by applying a three-day long wash-out period between tamoxifen treatment and toxin injection. Therefore, the Opn-iCreERT2 driver is best suited for the generation of mutant bile ducts, while the Ck19-CreERT driver has near absolute specificity for bile duct cells and is therefore favorable for lineage tracing experiments.

Highlights

The development of inducible cyclization recombinase (Cre) transgenic mouse strains has made it possible to generate cell type-specific knockout and lineage tracing experiments
Our results show that both Cre drivers nearly exclusively target cholangiocytes but that recombination in hepatocytes can occasionally occur in either model after tamoxifen injection, this is a rare event
This amounted to 1.80 independent recombination events per 100 cholangiocytes for Opn-CreERT -tdTomato mice and 1.91 for Cytokeratin 19 (Ck19)-CreERT -tdTomato mice observed at 8 weeks of age

Summary

Introduction

The development of inducible Cre transgenic mouse strains has made it possible to generate cell type-specific knockout and lineage tracing experiments. The second locus is a transgene or a knock-in where a cell type-specific promoter drives the expression of the Cre recombinase. CreER is released from the cell membrane and translocates into the nucleus where it catalyzes recombination between the two loxP sites. This results in the excision of a DNA segment that is flanked by two oriented loxP sites. This inducible CreER-loxP system is broadly used to conditionally knock out a gene of interest in specific cells by combining tissue specific CreER strains with tamoxifen injection at a specific time point. This conditional knock-out strategy prevents deletion of floxed segments prematurely or in non-desired cell types

Methods

Results

Conclusion