title: Generation of outbred Ace2 knockout mice by RNA transfection of TALENs displaying colitis reminiscent pathophysiology and inflammation.
abstract: The angiotensin I converting enzyme 2 (ACE2) is a key factor in the maintenance of intestinal homeostasis. Dysregulation of homeostasis can lead to inflammation of the colon (colitis), which can cause life-threatening enfeeblement or even cancer. Animal models are valuable surrogates in deciphering the pathology behind such human conditions and for screening of putative therapeutic targets or treatment paradigms. However, development of disease models can be time-consuming and technical demanding, which might hamper their application-value. In this study, we genetically disrupted the mouse Ace2 gene by direct injection of in vitro transcribed mRNA coding for transcription activator-like effector nucleases (TALENs) into the cytoplasm of outbred Kunming mouse zygotes. Consequently, somatic mutations were induced with an efficiency of 57%, of which 39% were frameshift mutations. Moreover, all modifications were stably transferred during germline transmission. In Ace2-knockout male mice (Ace2(-/y)), we observed severe chemical induced colitis, characterized by considerable weight loss, diarrhea and a shortened colon length. Histologically, Ace2 mutations resulted in the infiltration of leukocytes and the overt damage of the intestinal mucosal barrier. In addition, we detected an increased expression of inflammatory cytokines in the colon tissue of Ace2(-/y) mice. Collectively, the data indicate that high targeting efficiency and heritability can be achieved in an outbred mouse model by zygote injection of TALEN mRNA. Furthermore, the generated Ace2(-/y) mice display phenotypic traits reminiscent of colitis and we anticipate that such mice can be of value in studies of the intestinal microbiome or fecal transplantation.
authors.affiliation: BGI-Shenzhen, Beishan Industrial Zone, Yantian District, Shenzhen, 518083, China.
authors.forename: Chuxin
authors.initials: C
authors.lastname: Liu
authors.forename: Liping
authors.initials: L
authors.lastname: Xiao
authors.forename: Feida
authors.initials: F
authors.lastname: Li
authors.forename: Huanhuan
authors.initials: H
authors.lastname: Zhang
authors.forename: Qin
authors.initials: Q
authors.lastname: Li
authors.forename: Huan
authors.initials: H
authors.lastname: Liu
authors.forename: Shujin
authors.initials: S
authors.lastname: Fu
authors.forename: Chao
authors.initials: C
authors.lastname: Li
authors.forename: Xingju
authors.initials: X
authors.lastname: Zhang
authors.forename: Jun
authors.initials: J
authors.lastname: Wang
authors.forename: Nicklas H
authors.initials: NH
authors.lastname: Staunstrup
authors.forename: Yong
authors.initials: Y
authors.lastname: Li
authors.forename: Huanming
authors.initials: H
authors.lastname: Yang
mesh_terms: D000085962:Angiotensin-Converting Enzyme 2; D000818:Animals; D001483:Base Sequence; D003092:Colitis / Q000139:chemically induced / Q000235:genetics / Q000503:physiopathology*; D016207:Cytokines / Q000235:genetics / Q000378:metabolism; D016264:Dextran Sulfate / Q000633:toxicity; D004195:Disease Models, Animal; D004720:Endonucleases / Q000235:genetics; D005260:Female; D018095:Germ-Line Mutation; D008297:Male; D018345:Mice, Knockout; D008845:Microinjections; D008969:Molecular Sequence Data; D007703:Peptidyl-Dipeptidase A / Q000235:genetics*; D012333:RNA, Messenger; D014162:Transfection / Q000379:methods
chemical_list: D016207:Cytokines; D012333:RNA, Messenger; D016264:Dextran Sulfate; D004720:Endonucleases; D007703:Peptidyl-Dipeptidase A; C000705307:ACE2 protein, human; C000705328:Ace2 protein, mouse; D000085962:Angiotensin-Converting Enzyme 2