ABSTRACT
Preimplantation embryos are sensitive to maternal hormones affecting embryonic signal transduction and metabolic functions. We examined whether adiponectin, the most abundantly secreted adipokine, can influence glucose transport in mouse embryonic cells. In mouse blastocysts full-length adiponectin stimulated glucose uptake, while no effect of globular adiponectin was found. Full-length adiponectin stimulated translocation of GLUT8 glucose transporter to the cell membrane; we did not detect significant changes in the intracellular localization of GLUT4 glucose transporter in adiponectin-treated blastocysts. To study adiponectin signaling in detail, we used embryoid bodies formed from mouse embryonic carcinoma cell (ECC) line P19. We confirmed the expression of adiponectin receptors in these cells. Similar to mouse blastocysts, full-length adiponectin, but not globular adiponectin, stimulated glucose uptake in ECC P19 embryoid bodies. Moreover, full-length adiponectin stimulated AMPK and p38 MAPK phosphorylation. These results indicate that besides AMPK, p38 MAPK is a potential target of adiponectin in mouse embryonic cells. AMPK inhibitor did not influence the adiponectin-stimulated p38 MAPK phosphorylation, indicating independent action of these two signaling pathways. In mouse embryos adiponectin acts as a hormonal regulator of glucose uptake, which becomes especially important in phases with reduced levels of circulating insulin. Our results suggest that adiponectin maintains the glucose supply for early embryos under hypoinsulinaemic conditions, for example, in mothers suffering from type 1 diabetes mellitus.
Subject(s)
Adiponectin/physiology , Blastocyst/metabolism , Glucose Transport Proteins, Facilitative/metabolism , Glucose Transporter Type 4/metabolism , Glucose/metabolism , Animals , Cell Line, Tumor , Embryoid Bodies/metabolism , Female , MAP Kinase Signaling System , Mice , Receptors, Adiponectin/metabolismABSTRACT
Polo-like kinase 1 (PLK1), a member of the serine/threonine protein kinases family, is involved in multiple steps of mitotic progression. It regulates centrosome maturation, mitotic spindle formation, and cytokinesis. While studied extensively in somatic cells, little is known about PLK1 activities in the mammalian preimplantation embryo. We examined the role of PLK1 in the one-cell mouse embryo. Western blotting showed that the PLK1 protein content increased significantly during the S-phase of the one-cell stage and declined during the first mitotic division. Activation of PLK1 preceded nuclear envelope breakdown (NEBD) in both pronuclei at the entry to first embryo mitosis. Immunofluorescence revealed the presence of phosphorylated, active PLK1 (pThr(210) -PLK1) in both male and female pronuclei, and in the microtubule-organizing centers (MTOCs) shortly before NEBD. During the first mitotic metaphase, pThr(210) -PLK1 accumulated at the spindle poles and was also associated with condensed chromosomes. Inhibition of PLK1 activity with a specific PLK1 inhibitor, BI 2536, at the one-cell stage induced the formation of a bipolar spindle that displayed disordered microtubular arrangements and dislocated, condensed chromosomes. Although such embryos entered mitosis, they did not complete mitosis and arrested at metaphase. Time-lapse recording revealed progressive misalignment of condensed chromosomes during first mitotic metaphase. These data indicate that PLK1 activity is not essential for entry into first mitosis, but is required for the events leading up to metaphase-anaphase transition in the one-cell mouse embryo.
Subject(s)
Blastocyst/physiology , Cell Cycle Proteins/metabolism , Gene Expression Regulation, Developmental/physiology , Mitosis/physiology , Protein Serine-Threonine Kinases/metabolism , Proto-Oncogene Proteins/metabolism , Animals , Blastocyst/metabolism , Blotting, Western , Cell Cycle Proteins/antagonists & inhibitors , Female , Fluorescent Antibody Technique , Gene Expression Regulation, Developmental/genetics , Male , Mice , Microtubule-Organizing Center/metabolism , Protein Serine-Threonine Kinases/antagonists & inhibitors , Proto-Oncogene Proteins/antagonists & inhibitors , Pteridines/pharmacology , Spindle Apparatus/drug effects , Spindle Apparatus/physiology , Time-Lapse Imaging , Polo-Like Kinase 1ABSTRACT
S. typhimurium isolates obtained during a large outbreak of human salmonellosis associated with smoked mackerels in the Czech Republic as well as strains of S. typhimurium isolated from black headed gull (Larus ridibundus) were examined following biotyping, phage typing, plasmid profiling and restriction endonuclease analysis (Eco RI, Hind III and Bam HI) of plasmid DNA. The epidemic strain of S. typhimurium and two isolates from environment of nesting colony black-headed gull were meso-inositol and L-rhamnose negative, phage type 141. The isolates from human and environment of nesting colony were found to share the same plasmid profile and REA.
