title: Functional role of arginine during the peri-implantation period of pregnancy. II. Consequences of loss of function of nitric oxide synthase NOS3 mRNA in ovine conceptus trophectoderm.
abstract: Nitric oxide (NO) is a gaseous molecule that regulates angiogenesis and vasodilation via activation of the cGMP pathway. However, functional roles of NO during embryonic development from spherical blastocysts to elongated filamentous conceptuses (embryo and extraembryonic membrane) during the peri-implantation period of pregnancy have not been elucidated in vivo. In order to assess roles of NO production in survival and development of the ovine conceptus, we conducted an in vivo morpholino antisense oligonucleotide (MAO)-mediated knockdown trial of nitric oxide synthase-3 (NOS3) mRNA, the major isoform of NO synthase, in ovine conceptus trophectoderm (Tr). Translational knockdown of NOS3 mRNA results in small, thin, and underdeveloped conceptuses, but normal production of interferon-tau, the pregnancy recognition signal in sheep. MAO-NOS3 knockdown in conceptuses decreased the abundance of NOS3 (72%, P < 0.05) and the arginine transporter SLC7A1 proteins in conceptus Tr. Furthermore, the amounts of ornithine and polyamines were less (P < 0.01) in uterine fluid, whereas the amounts of arginine (58%, P < 0.01), citrulline (68%, P < 0.05), ornithine (68%, P < 0.001), glutamine (78%, P < 0.001), glutamate (68%, P < 0.05), and polyamines (P < 0.01) were less in conceptuses, which likely accounts for the failure of MAO-NOS3 conceptuses to develop normally. For MAO-NOS3 conceptuses, there were no compensatory increases in the expression levels of either nitric oxide synthase-1 (NOS1) or nitric oxide synthase-2 (NOS2) or in expression of enzymes for synthesis of polyamines (ornithine decarboxylase, arginine decarboxylase, agmatinase) from arginine or ornithine with which to rescue development of MAO-NOS3 conceptuses. Thus, the adverse effect of MAO-NOS3 to reduce NO generation and the transport of arginine and ornithine into conceptuses is central to an explanation for failure of normal development of MAO-NOS3, compared to control conceptuses. The study, for the first time, created an NO-deficient mammalian conceptus model in vivo and provided new insights into the orchestrated events of conceptus development during the peri-implantation period of pregnancy. Our data suggest that NOS3 is the key enzyme for NO production by conceptus Tr and that this protein also regulates the availability of arginine in conceptus tissues for synthesis of polyamines that are essential for conceptus survival and development.
authors: Center for Animal Biotechnology and Genomics, Texas A&M University, College Station, Texas Department of Animal Science, Texas A&M University, College Station, Texas.
authors: Xiaoqiu
authors: X
authors: Wang
authors: Department of Veterinary Integrative Biosciences, Texas A&M University, College Station, Texas.
authors: James W
authors: JW
authors: Frank
authors: Department of Animal Science, Texas A&M University, College Station, Texas.
authors: Jing
authors: J
authors: Xu
authors: Department of Animal Science, Texas A&M University, College Station, Texas.
authors: Kathrin A
authors: KA
authors: Dunlap
authors: Department of Animal Science, Texas A&M University, College Station, Texas.
authors: M Carey
authors: MC
authors: Satterfield
authors: Department of Veterinary Integrative Biosciences, Texas A&M University, College Station, Texas.
authors: Robert C
authors: RC
authors: Burghardt
authors: Department of Biomedical Sciences, Colorado State University, Fort Collins, Colorado.
authors: Jared J
authors: JJ
authors: Romero
authors: Department of Biomedical Sciences, Colorado State University, Fort Collins, Colorado.
authors: Thomas R
authors: TR
authors: Hansen
authors: Center for Animal Biotechnology and Genomics, Texas A&M University, College Station, Texas Department of Animal Science, Texas A&M University, College Station, Texas.
authors: Guoyao
authors: G
authors: Wu
authors: Center for Animal Biotechnology and Genomics, Texas A&M University, College Station, Texas Department of Animal Science, Texas A&M University, College Station, Texas fbazer@cvm.tamu.edu.
authors: Fuller W
authors: FW
authors: Bazer
mesh_terms: D000818:Animals; D017849:Animals, Inbred Strains; D001120:Arginine / Q000378:metabolism*; D001755:Blastocyst / Q000166:cytology / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology; D027201:Cationic Amino Acid Transporter 1 / Q000378:metabolism; D010064:Embryo Implantation / Q000187:drug effects; D004622:Embryo, Mammalian / Q000166:cytology / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology; D047108:Embryonic Development / Q000187:drug effects; D005321:Extraembryonic Membranes / Q000166:cytology / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology; D005260:Female; D005317:Fetal Growth Retardation / Q000139:chemically induced / Q000378:metabolism / Q000473:pathology; D007150:Immunohistochemistry; D007370:Interferon Type I / Q000378:metabolism; D060172:Morpholinos / Q000494:pharmacology; D052250:Nitric Oxide Synthase Type III / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D011247:Pregnancy; D011255:Pregnancy Maintenance / Q000187:drug effects; D011257:Pregnancy Proteins / Q000378:metabolism; D012333:RNA, Messenger / Q000037:antagonists & inhibitors / Q000378:metabolism*; D034561:Sheep, Domestic; D013781:Texas
chemical_list: D027201:Cationic Amino Acid Transporter 1; D007370:Interferon Type I; D060172:Morpholinos; D011257:Pregnancy Proteins; D012333:RNA, Messenger; C020947:interferon tau; D001120:Arginine; D052250:Nitric Oxide Synthase Type III
keywords: development; morpholino; nitric oxide; nitric oxide synthase