title: Vascular endothelium-specific overexpression of human catalase in cloned pigs.
abstract: The objective of this study was to develop transgenic Yucatan minipigs that overexpress human catalase (hCat) in an endothelial-specific manner. Catalase metabolizes hydrogen peroxide (H(2)O(2)), an important regulator of vascular tone that contributes to diseases such as atherosclerosis and preeclampsia. A large animal model to study reduced endothelium-derived H(2)O(2) would therefore generate valuable translational data on vascular regulation in health and disease. Yucatan minipig fetal fibroblasts stably co-transfected with human catalase (Tie2-hCat) and eGFP expression constructs were isolated into single-cell populations. The presence of the Tie2-hCat transgene in individual colonies of fibroblasts was determined by PCR. Transgenic fibroblasts were used for nuclear transfer into enucleated oocytes by electrofusion. A minimum of 140 cloned embryos were transferred per surrogate sow (n = 4). All four surrogates maintained pregnancies and piglets were delivered by cesarean section. Nine male piglets from three of the four litters carried the Tie2-hCat transgene. Expression of human catalase mRNA and overall elevated catalase protein in isolated umbilical endothelial cells from transgenic piglets were verified by RT-PCR and western blot, respectively, and endothelial localization was confirmed by immunohistochemistry. Increased enzymatic activity of catalase in transgenic versus wild-type endothelial cells was inferred based on significantly reduced levels of H(2)O(2) in culture. The similarities in swine and human cardiovascular anatomy and physiology will make this pig model a valuable source of information on the putative role of endothelium-derived H(2)O(2) in vasodilation and in the mechanisms underlying vascular health and disease.
authors.affiliation: Department of Biomedical Sciences, University of Missouri, Columbia, MO 65211, USA. whytej@missouri.edu
authors.forename: J J
authors.initials: JJ
authors.lastname: Whyte
authors.forename: M
authors.initials: M
authors.lastname: Samuel
authors.forename: E
authors.initials: E
authors.lastname: Mahan
authors.forename: J
authors.initials: J
authors.lastname: Padilla
authors.forename: G H
authors.initials: GH
authors.lastname: Simmons
authors.forename: A A
authors.initials: AA
authors.lastname: Arce-Esquivel
authors.forename: S B
authors.initials: SB
authors.lastname: Bender
authors.forename: K M
authors.initials: KM
authors.lastname: Whitworth
authors.forename: Y H
authors.initials: YH
authors.lastname: Hao
authors.forename: C N
authors.initials: CN
authors.lastname: Murphy
authors.forename: E M
authors.initials: EM
authors.lastname: Walters
authors.forename: R S
authors.initials: RS
authors.lastname: Prather
authors.forename: M H
authors.initials: MH
authors.lastname: Laughlin
mesh_terms: D000818:Animals; D030801:Animals, Genetically Modified; D002318:Cardiovascular Diseases / Q000201:enzymology / Q000235:genetics; D002374:Catalase / Q000235:genetics* / Q000378:metabolism; D019976:Cloning, Organism; D004195:Disease Models, Animal; D004624:Embryo Transfer; D004730:Endothelium, Vascular / Q000166:cytology / Q000378:metabolism; D005260:Female; D015870:Gene Expression; D006801:Humans; D006861:Hydrogen Peroxide / Q000378:metabolism*; D008297:Male; D011247:Pregnancy; D042787:Receptor, TIE-2 / Q000235:genetics; D013552:Swine; D013556:Swine, Miniature / Q000235:genetics* / Q000378:metabolism
chemical_list: D006861:Hydrogen Peroxide; D002374:Catalase; D042787:Receptor, TIE-2