title: Cellular characterization of thrombocytes in Xenopus laevis with specific monoclonal antibodies.
abstract: Platelets are produced from megakaryocytes (MKs) in the bone marrow. In contrast, most nonmammalian vertebrates have nucleated and spindle-shaped thrombocytes instead of platelets in their circulatory systems, and the presence of MKs as thrombocyte progenitors has not been verified. In developing a new animal model in adult African clawed frog (Xenopus laevis), we needed to distinguish nucleated thrombocytes and their progenitors from other blood cells, because the cellular morphology of activated thrombocytes resembles lymphocytes and other cells. We initially generated two monoclonal antibodies, T5 and T12, to X. laevis thrombocytes. Whereas T5 recognized both thrombocytes and leukocytes, T12 specifically reacted to spindle-shaped thrombocytes. The T12(+) thrombocytes displayed much higher DNA ploidy than nucleated erythrocytes, and they expressed CD41 and Fli-1. In the presence of CaCl2, adenosine diphosphate, thrombin, or various collagens, T12(+) thrombocytes exhibited aggregation. These thrombocytes were located predominantly in the hepatic sinusoids and the splenic red pulp, suggesting that both organs are the sites of thrombopoiesis. Notably, circulating thrombocytes exhibited lower DNA ploidy than hepatic thrombocytes. Intraperitoneal administration of T12 produced immune thrombocytopenia in frogs, which reached a nadir 4 days postinjection, followed by recovery, suggesting that humoral regulation maintained the number of circulating thrombocytes. Although differences between MKs and thrombocytes in X. laevis remain to be defined, our results provide further insight into MK development and thrombopoiesis in vertebrates.
authors.affiliation: Integrative Bioscience and Biomedical Engineering, Graduate School of Advanced Science and Engineering, Waseda University, Tokyo, Japan; Research Fellow of the Japan Society for the Promotion of Science, Tokyo, Japan.
authors.forename: Yuta
authors.initials: Y
authors.lastname: Tanizaki
authors.affiliation: Department of Biology, Education and Integrated Arts and Sciences, Waseda University, Tokyo, Japan.
authors.forename: Takako
authors.initials: T
authors.lastname: Ishida-Iwata
authors.affiliation: Department of Biology, Education and Integrated Arts and Sciences, Waseda University, Tokyo, Japan.
authors.forename: Miyako
authors.initials: M
authors.lastname: Obuchi-Shimoji
authors.affiliation: Integrative Bioscience and Biomedical Engineering, Graduate School of Advanced Science and Engineering, Waseda University, Tokyo, Japan; Department of Biology, Education and Integrated Arts and Sciences, Waseda University, Tokyo, Japan. Electronic address: tkato@waseda.jp.
authors.forename: Takashi
authors.initials: T
authors.lastname: Kato
mesh_terms: D000244:Adenosine Diphosphate / Q000494:pharmacology; D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis / Q000494:pharmacology*; D015415:Biomarkers / Q000378:metabolism; D001792:Blood Platelets / Q000166:cytology* / Q000187:drug effects / Q000276:immunology; D002122:Calcium Chloride / Q000494:pharmacology; D002478:Cells, Cultured; D003094:Collagen / Q000494:pharmacology; D015870:Gene Expression; D008099:Liver / Q000166:cytology* / Q000187:drug effects / Q000276:immunology; D008214:Lymphocytes / Q000166:cytology / Q000187:drug effects / Q000276:immunology; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D009928:Organ Specificity; D010974:Platelet Aggregation; D040201:Platelet Membrane Glycoprotein IIb / Q000235:genetics / Q000276:immunology; D011003:Ploidies; D051789:Proto-Oncogene Protein c-fli-1 / Q000235:genetics / Q000276:immunology; D013154:Spleen / Q000166:cytology* / Q000187:drug effects / Q000276:immunology; D013917:Thrombin / Q000494:pharmacology; D013921:Thrombocytopenia / Q000139:chemically induced / Q000276:immunology / Q000473:pathology; D034061:Thrombopoiesis; D014982:Xenopus laevis / Q000235:genetics / Q000276:immunology
chemical_list: D000911:Antibodies, Monoclonal; D015415:Biomarkers; D040201:Platelet Membrane Glycoprotein IIb; D051789:Proto-Oncogene Protein c-fli-1; D000244:Adenosine Diphosphate; D003094:Collagen; D013917:Thrombin; D002122:Calcium Chloride