Generation of heterozygous fibrillin-1 mutant cloned pigs from genome-edited foetal fibroblasts

Umeyama, Kazuhiro; Watanabe, Kota; Watanabe, Masaru; Horiuchi, Keisuke; Nakano, K.; Kitashiro, Masateru; Matsunari, Hitomi; Kimura, Tokuhiro; Arima, Yoshimi; Sampetrean, Oltea; Nagaya, Masaki; Saito, Masahiro; Saya, Hideyuki; Kosaki, Kenjiro; Nagashima, Hiroshi; Matsumoto, Morio

doi:10.1038/srep24413

Cited by 35 publications

(39 citation statements)

References 45 publications

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“…However, all of the cloned offspring were stillborn. Because the body weights of the stillborn piglets were similar to the average weight of the previously cloned piglets [25, 34], death may have occurred as a result of an intrapartum accident. Overexpression of mPlum might also have manifested cytotoxicity.…”

Section: Discussionmentioning

confidence: 99%

A transgenic-cloned pig model expressing non-fluorescent modified Plum

Nagaya

Watanabe

Kobayashi

et al. 2016

Journal of Reproduction and Development

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Genetically modified pigs that express fluorescent proteins such as green and red fluorescent proteins have become indispensable biomedical research tools in recent years. Cell or tissue transplantation studies using fluorescent markers should be conducted, wherein the xeno-antigenicity of the fluorescent proteins does not affect engraftment or graft survival. Thus, we aimed to create a transgenic (Tg)-cloned pig that was immunologically tolerant to fluorescent protein antigens. In the present study, we generated a Tg-cloned pig harboring a derivative of Plum modified by a single amino acid substitution in the chromophore. The cells and tissues of this Tg-cloned pig expressing the modified Plum (mPlum) did not fluoresce. However, western blot and immunohistochemistry analyses clearly showed that the mPlum had the same antigenicity as Plum. Thus, we have obtained primary proof of principle for creating a cloned pig that is immunologically tolerant to fluorescent protein antigens.

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Section: Discussionmentioning

confidence: 99%

A transgenic-cloned pig model expressing non-fluorescent modified Plum

Nagaya

Watanabe

Kobayashi

et al. 2016

Journal of Reproduction and Development

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“…Genome-editing technologies have been used to generate animal models to study cardiomyopathy [69][70][71][72][73]. One of the most promising approaches entails the use of somatic in vivo genome editing, in which genome editing tools are delivered into a target organ in an adult animal.…”

Section: In Vivo Modelsmentioning

confidence: 99%

Genome Editing for the Understanding and Treatment of Cardiomyopathies

Nguyen¹,

Lim²,

Yokota³

2019

Preprint

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Cardiomyopathies are diseases of heart muscle, a significant percentage of which are genetic in origin. Cardiomyopathies can be classified as dilated, hypertrophic, restrictive, arrhythmogenic right ventricular or left ventricular non-compaction, although mixed morphologies are possible. A subset of neuromuscular disorders, notably Duchenne and Becker muscular dystrophies, are also characterized by cardiomyopathy aside from skeletal myopathy. The global burden of cardiomyopathies is certainly high, necessitating further research and novel therapies. Genome editing tools, which include zinc finger nucleases (ZFNs), transcription activator-like effector nucleases (TALENs) and clustered regularly interspaced short palindromic repeats (CRISPR) systems have emerged as increasingly important technologies in studying this group of cardiovascular disorders. In this review, we discuss the applications of genome editing in the understanding and treatment of cardiomyopathy. We also describe recent advances in genome editing that may help improve these applications, and some future prospects for genome editing in cardiomyopathy treatment.Keywords: dilated cardiomyopathy (DCM); hypertrophic cardiomyopathy (HCM); restrictive cardiomyopathy (RCM); arrhythmogenic right ventricular cardiomyopathy (ARVC); left ventricular non-compaction cardiomyopathy (LVNC); Duchenne muscular dystrophy; dystrophin; genome editing; CRISPR/Cas9; Cpf1 (Cas12a) Cardiomyopathy is also a major manifestation in a subset of neuromuscular disorders [7,8]. Cardiomyopathy is most commonly associated with Duchenne muscular dystrophy (DMD) and Becker muscular dystrophy (BMD), both X-linked recessive disorders caused by mutations in the dystrophin (DMD) gene [8]. DMD codes for dystrophin, a cytoskeletal protein that maintains the Preprints (www.preprints.org) | NOT PEER-REVIEWED | Posted: structural integrity of muscle fibres during contraction-relaxation cycles. Loss-of-function mutations in the DMD gene result in an absence of dystrophin, leading to DMD [9][10][11]. Mutations maintaining the DMD reading frame produce truncated but partly functional dystrophin, and often result in a milder form of the disease known as BMD [12,13]. Cardiomyopathy is a leading cause of death in both DMD and BMD patients and is routinely described as being DCM [13,14]. Other neuromuscular disorders associated with cardiomyopathy include the limb girdle muscle dystrophies, myotonic dystrophy, and Friedreich ataxia [15].Advances in molecular genetics enable genome editing to be incorporated into various fields of biomedical research, including the cardiovascular sciences. Currently, the most commonly used tools are zinc finger nucleases (ZFNs), transcription activator-like effector nucleases (TALENs), and clustered regularly interspaced short palindromic repeats (CRISPR) [16]. Both ZFNs and TALENs are engineered restriction enzymes created by combining a DNA binding domain with a DNA cleavage domain adapted from the FokI restriction enzyme [17][18][19]. The DNA binding domain in Z...

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“…If a conditional knockout system is preferred, loxP sites flanking the target gene can be introduced using repair templates [54]. To date, CVD has been studied in a variety of generated mammalian models, including but not limited to rats [55–60], rabbits [61–63], and pigs [64, 65]. In zebrafish models alone, cardiac development [66, 67], cardiac regeneration [68], vascular development [69, 70], and inherited cardiomyopathy [71] have been explored.…”

Section: Using Genome Editing To Create In Vivo Disease Modelsmentioning

confidence: 99%

Genome Editing for the Study of Cardiovascular Diseases

Chadwick

Musunuru

2017

Curr Cardiol Rep

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Purpose of Review The opportunities afforded through the recent advent of genome-editing technologies have allowed investigators to more easily study a number of diseases. The advantages and limitations of the most prominent genome-editing technologies are described in this review, along with potential applications specifically focused on cardiovascular diseases. Recent Findings The recent genome-editing tools using programmable nucleases, such as zinc-finger nucleases, transcription activator-like effector nucleases, and clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated 9 (Cas9), have rapidly been adapted to manipulate genes in a variety of cellular and animal models. A number of recent cardiovascular disease-related publications report cases in which specific mutations are introduced into disease models for functional characterization and for testing of therapeutic strategies. Summary Recent advances in genome-editing technologies offer new approaches to understand and treat diseases. Here, we discuss genome editing strategies to easily characterize naturally occurring mutations and offer strategies with potential clinical relevance.

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Generation of heterozygous fibrillin-1 mutant cloned pigs from genome-edited foetal fibroblasts

Cited by 35 publications

References 45 publications

A transgenic-cloned pig model expressing non-fluorescent modified Plum

A transgenic-cloned pig model expressing non-fluorescent modified Plum

Genome Editing for the Understanding and Treatment of Cardiomyopathies

Genome Editing for the Study of Cardiovascular Diseases

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