14
views
0
recommends
+1 Recommend
0 collections
    0
    shares
      • Record: found
      • Abstract: found
      • Article: found
      Is Open Access

      Generation of transgenic golden Syrian hamsters

      letter

      Read this article at

      Bookmark
          There is no author summary for this article yet. Authors can add summaries to their articles on ScienceOpen to make them more accessible to a non-specialist audience.

          Abstract

          Dear Editor, Golden Syrian hamsters are small rodents, but they display many features that resemble the physiology and metabolism of humans. Hamsters have been widely used in many research areas, including carcinogenesis 1 , reproduction 2 , virology 3 , diabetes 4 and cardiovascular diseases 5 . With respect to lipid and glucose metabolism, hamsters, like humans, exhibit high levels of cholesteryl ester transport protein (CETP), intestinal-only ApoB editing, low levels of hepatic low-density lipoprotein (LDL) receptor activity 6 and a high glycemic response to dietary fructose 7 , all of which are not observed in other rodents such as mice and rats. Consequently, hamsters, like humans, exhibit enhanced susceptibility to atherosclerosis (AS) and diabetes 8 , which led to the widespread use of hamsters in studies on AS and diabetes. In the past 2-3 decades, due to the fast development of transgenic and knockout mice, hamsters were gradually replaced by these mouse models. However, due to multiple differences between mice and humans with respect to physiology and metabolism, the use of gene-manipulated mice has limited value in disease modeling and pathophysiological studies. Extensive literature search has revealed an absence of reports on genetically manipulated hamster models. To capitalize on the special metabolic features of hamsters, we aim to generate gene-manipulated hamsters as an alternate rodent model for general applications. As the initial step to create a genetically manipulated hamster, we utilized a highly efficient lentiviral vector to generate transgenic hamsters expressing enhanced green fluorescent protein (eGFP). By modifying and optimizing the protocols for producing transgenic mice and rabbits in our laboratory 9,10 , we developed a specific procedure for hamster superovulation, fertilized egg harvesting, perivitelline space microinjection and embryo transfer. After the successful culture of fertilized hamster eggs that developed into 4- and 8-cell embryos in vitro (Figure 1A), we implanted these embryos into pseudopregnant females. We obtained 7-10 pups/litter in 4 out of 7 surrogate mothers. Next, we microinjected 50-100 picoliters of a lentiviral eGFP vector (Figure 1B) at a titer of 2 × 109 titer units/ml into the perivitelline space of the fertilized eggs to generate transgenic hamsters that express eGFP. A total of 6 out of 32 live-born pups from 5 surrogate mothers each receiving 30-40 microinjected eggs were identified as being eGFP-positive by PCR genotyping, and 5 of the pups were further validated by Southern blot analysis (Figure 1C and 1D). Of the 5 positive pups validated by Southern blotting, 2 of them (males) expressed eGFP in the exposed skin area as determined by direct fluorescence imaging (Figure 1E). Transgenic lines were then established by breeding the 2 founders (F0) with non-transgenic females. Among the progenies from 3 litters, 21% of the animals were eGFP-positive as determined by direct fluorescence imaging. Representative fluorescence images of the first generation (F1) pups from one of the 3 litters are shown in Figure 1F. All the examined organs from one eGFP-positive F1 pup showed strong-to-moderate levels of green fluorescence, which include the liver, kidney, heart, skeletal muscle, lung, brain, white/brown adipose tissues, adrenal glands and eyes; however, no fluorescence was observed in these organs of the littermate control (Figure 1G and data not shown). The peritoneal macrophages and bone marrow cells from another eGFP-transgenic F1 hamster also demonstrated an approximately 70% eGFP-positive cell population as analyzed by FACS (Figure 1H). With the establishment of transgenic hamster model in the present study, it is now possible to generate small rodent disease models that recapitulate human pathogenesis. For example, generation of a transgenic hamster overexpressing proprotein convertase subtilisin/kexin type 9 (PCSK9), a protein involved in cholesterol homeostasis by inducing LDL receptor degradation, will be highly desirable. Although mice and pigs overexpressing PCSK9 developed hypercholesterolemia and AS 11 , these animals, unlike humans, do not express CETP, a critical factor involved in lipid transport and AS development. Along with the recent development of genome-editing methods such as TALENs (transcription activator-like effector nucleases) and the CRISPR/Cas system (clustered regularly interspaced short palindromic repeats/CRISPR-associated system) 12 , the generation of gene-targeted hamster models is thus warranted in conjunction with the hamster embryo manipulation method optimized in the present study. These hamster models will be utilized extensively in studies for metabolic cardiovascular diseases and will become the candidate models when genomics and proteomics tools are fully developed for this species in the future. Genetically altered hamsters may potentially replace mice as the mainstream animal model for metabolic cardiovascular research. Therefore, the present study will promote the use of genetically engineered hamsters as disease models. These models might recapitulate many features of human metabolic disorders while simultaneously retaining the ease of handling, the simplicity of management and the cost efficiency of small rodents as compared to genetically engineered large animals such as mini-pigs 13 or non-human primates 14 . Detailed methods are described in the Supplementary information, Data S1.

          Related collections

          Author and article information

          Journal
          Cell Res
          Cell Res
          Cell Research
          Nature Publishing Group
          1001-0602
          1748-7838
          March 2014
          07 January 2014
          1 March 2014
          : 24
          : 3
          : 380-382
          Affiliations
          [1 ]Institute of Cardiovascular Sciences and Key Laboratory of Molecular Cardiovascular Sciences, Ministry of Education, Peking University , Beijing 100191, China
          [2 ]Department of Endocrinology, Lu He Teaching Hospital of the Capital Medical University , Beijing 101149, China
          [3 ]Center for Molecular Biology Laboratory and Department of Animal Science, Xinyang College of Agriculture and Forestry , Xinyang Yangshan new district 24 new street, Xinyang, Henan 464000, China
          [4 ]Department of Animal and Avian Sciences, University of Maryland , College Park, MD 20742, USA
          Author notes
          [*]

          These two authors contributed equally to this work.

          Article
          cr20142
          10.1038/cr.2014.2
          3945892
          24394888
          ff92d310-ad3f-49b0-aae3-42db748d6441
          Copyright © 2014 Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences

          This work is licensed under the Creative Commons Attribution-NonCommercial-No Derivative Works 3.0 Unported License. To view a copy of this license, visit http://creativecommons.org/licenses/by-nc-nd/3.0

          History
          Categories
          Letter to the Editor

          Cell biology
          Cell biology

          Comments

          Comment on this article