Establishment of a Cre-rat resource for creating  conditional and physiological relevant models of human diseases

Zhang, Huimin; Zheng, Qi; Chen-Tsai, Ruby Yanru

doi:10.1007/s11248-020-00226-7

Cited by 10 publications

(9 citation statements)

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“…The Cre/ loxP system is extensively utilized for spatial or temporal regulation of gene function in rats and mice [ 38 , 39 , 40 , 41 ]. In vivo observation of cell fusion or transdifferentiation by using the Cre/ loxP system has been performed and the Cre/ loxP system is regarded as a useful way [ 42 , 43 , 44 ].…”

Section: Discussionmentioning

confidence: 99%

Differentiation Capacity of Bone Marrow-Derived Rat Mesenchymal Stem Cells from DsRed and Cre Transgenic Cre/loxP Models

Tseng

Lee

et al. 2022

Cells

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Cre/loxP recombination is a well-established technique increasingly used for modifying DNA both in vitro and in vivo. Nucleotide alterations can be edited in the genomes of mammalian cells, and genetic switches can be designed to target the expression or excision of a gene in any tissue at any time in animal models. In this study, we propose a system which worked via the Cre/loxP switch gene and DsRed/emGFP dual-color fluorescence imaging. Mesenchymal stem cells (MSCs) can be used to regenerate damaged tissue because of their differentiation capacity. Although previous studies have presented evidence of fusion of transplanted MSCs with recipient cells, the possibility of fusion in such cases remains debated. Moreover, the effects and biological implications of the fusion of MSCs at the tissue and organ level have not yet been elucidated. Thus, the method for determining this issue is significant and the models we proposed can illustrate the question. However, the transgenic rats exhibited growth slower than that of wild-type rats over several weeks. The effects on the stemness, proliferation, cell cycle, and differentiation ability of bone marrow–derived rat MSCs (BM-rMSCs) from the models were examined to ensure our design was appropriate for the in vivo application. We demonstrated that MSC surface markers were maintained in DsRed and Cre transgenic rMSCs (DsRed-rMSCs and Cre-rMSCs, respectively). A WST-8 assay revealed decreased proliferative activity in these DsRed-rMSCs and Cre-rMSCs; this result was validated through cell counting. Furthermore, cell cycle analysis indicated a decrease in the proportion of G1-phase cells and a concomitant increase in the proportion of S-phase cells. The levels of cell cycle–related proteins also decreased in the DsRed-rMSCs and Cre-rMSCs, implying decelerated phase transition. However, the BM-rMSCs collected from the transgenic rats did not exhibit altered adipogenesis, osteogenesis, or chondrogenesis. The specific markers of these types of differentiation were upregulated after induction. Therefore, BM-rMSCs from DsRed and Cre transgenic models can be used to investigate the behavior of MSCs and related mechanisms. Such application may further the development of stem cell therapy for tissue damage and other diseases.

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Section: Discussionmentioning

confidence: 99%

Differentiation Capacity of Bone Marrow-Derived Rat Mesenchymal Stem Cells from DsRed and Cre Transgenic Cre/loxP Models

Tseng

Lee

et al. 2022

Cells

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“…If the RNP enters the nuclei of cells, GE should commence quickly, which will, in turn, lead to increased GE efficiency. Using this RNP-based GE system, a number of GE rats have been produced to date [ 57 , 58 , 68 , 90 , 91 , 95 , 111 , 113 , 115 , 117 , 121 , 126 , 131 , 142 , 150 ].…”

Section: Delivery Methodsmentioning

confidence: 99%

Recent Advances in the Production of Genome-Edited Rats

Nakamura

Inada

Takabayashi

2022

IJMS

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The rat is an important animal model for understanding gene function and developing human disease models. Knocking out a gene function in rats was difficult until recently, when a series of genome editing (GE) technologies, including zinc-finger nucleases (ZFNs), transcription activator-like effector nucleases (TALENs), and the type II bacterial clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated Cas9 (CRISPR/Cas9) systems were successfully applied for gene modification (as exemplified by gene-specific knockout and knock-in) in the endogenous target genes of various organisms including rats. Owing to its simple application for gene modification and its ease of use, the CRISPR/Cas9 system is now commonly used worldwide. The most important aspect of this process is the selection of the method used to deliver GE components to rat embryos. In earlier stages, the microinjection (MI) of GE components into the cytoplasm and/or nuclei of a zygote was frequently employed. However, this method is associated with the use of an expensive manipulator system, the skills required to operate it, and the egg transfer (ET) of MI-treated embryos to recipient females for further development. In vitro electroporation (EP) of zygotes is next recognized as a simple and rapid method to introduce GE components to produce GE animals. Furthermore, in vitro transduction of rat embryos with adeno-associated viruses is potentially effective for obtaining GE rats. However, these two approaches also require ET. The use of gene-engineered embryonic stem cells or spermatogonial stem cells appears to be of interest to obtain GE rats; however, the procedure itself is difficult and laborious. Genome-editing via oviductal nucleic acids delivery (GONAD) (or improved GONAD (i-GONAD)) is a novel method allowing for the in situ production of GE zygotes existing within the oviductal lumen. This can be performed by the simple intraoviductal injection of GE components and subsequent in vivo EP toward the injected oviducts and does not require ET. In this review, we describe the development of various approaches for producing GE rats together with an assessment of their technical advantages and limitations, and present new GE-related technologies and current achievements using those rats in relation to human diseases.

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“…The RRRC also provides colony management and breeding services, genetic testing, and microinjection services. Recently, a total of ten Cre rat lines were generated, including eight for neural-specific and two for cardiovascular-specific Cre expression [ 10 ].…”

Section: Rat Genome Resourcesmentioning

confidence: 99%

Positional cloning of rat mutant genes reveals new functions of these genes

Kuramoto

2023

Exp. Anim.

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The laboratory rat (Rattus norvegicus) is a key model organism for biomedical research. Rats can be subjected to strict genetic and environmental controls. The rat's large body size is suitable for both surgical operations and repeated measurements of physiological parameters. These advantages have led to the development of numerous rat models for genetic diseases. Forward genetics is a proven approach for identifying the causative genes of these disease models but requires genome resources including genetic markers and genome sequences. Over the last few decades, rat genome resources have been developed and deposited in bioresource centers, which have enabled us to perform positional cloning in rats. To date, more than 100 disease-related genes have been identified by positional cloning. Since some disease models are more accessible in rats than mice, the identification of causative genes in these models has sometimes led to the discovery of novel functions of genes. As before, various mutant rats are also expected to be discovered and developed as disease models in the future. Thus, the forward genetics continues to be an important approach to find genes involved in disease phenotypes in rats. In this review, I provide an overview the development of rat genome resources and describe examples of positional cloning in rats in which novel gene functions have been identified. Rat Resource and Research Center (RRRC)The Rat Resource and Research Center (RRRC, http://www.rrrc.us/) is managed by the University of Missouri. The RRRC's major services are to curate valuable rat lines for cryopreservation and rederivation, as well as to distribute rat strains. The Center's research is focused on improving cryopreservation and in vitro fertilization, generating new rat models, and characterizing the gut microbiota of rat models. The RRRC also provides colony management and breeding services, genetic testing, and microinjection services. Recently, a total of ten Cre rat lines were generated, including eight for neural-specific and two for cardiovascular-specific Cre expression [10].

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Establishment of a Cre-rat resource for creating conditional and physiological relevant models of human diseases

Cited by 10 publications

References 50 publications

Differentiation Capacity of Bone Marrow-Derived Rat Mesenchymal Stem Cells from DsRed and Cre Transgenic Cre/loxP Models

Differentiation Capacity of Bone Marrow-Derived Rat Mesenchymal Stem Cells from DsRed and Cre Transgenic Cre/loxP Models

Recent Advances in the Production of Genome-Edited Rats

Positional cloning of rat mutant genes reveals new functions of these genes

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