title: The expression of embryonic globin mRNA in a severely anemic mouse model induced by treatment with nitrogen-containing bisphosphonate.
abstract: BACKGROUND
  Mammalian erythropoiesis can be divided into two distinct types, primitive and definitive, in which new cells are derived from the yolk sac and hematopoietic stem cells, respectively. Primitive erythropoiesis occurs within a restricted period during embryogenesis. Primitive erythrocytes remain nucleated, and their hemoglobins are different from those in definitive erythrocytes. Embryonic type hemoglobin is expressed in adult animals under genetically abnormal condition, but its later expression has not been reported in genetically normal adult animals, even under anemic conditions. We previously reported that injecting animals with nitrogen-containing bisphosphonate (NBP) decreased erythropoiesis in bone marrow (BM). Here, we induced severe anemia in a mouse model by injecting NBP injection in combination with phenylhydrazine (PHZ), and then we analyzed erythropoiesis and the levels of different types of hemoglobin.
  
  
  METHODS
  Splenectomized mice were treated with NBP to inhibit erythropoiesis in BM, and with PHZ to induce hemolytic anemia. We analyzed hematopoietic sites and peripheral blood using morphological and molecular biological methods.
  
  
  RESULTS
  Combined treatment of splenectomized mice with NBP and PHZ induced critical anemia compared to treatment with PHZ alone, and numerous nucleated erythrocytes appeared in the peripheral blood. In the BM, immature CD71-positive erythroblasts were increased, and extramedullary erythropoiesis occurred in the liver. Furthermore, embryonic type globin mRNA was detected in both the BM and the liver. In peripheral blood, spots that did not correspond to control hemoglobin were observed in 2D electrophoresis. ChIP analyses showed that KLF1 and KLF2 bind to the promoter regions of β-like globin. Wine-colored capsuled structures were unexpectedly observed in the abdominal cavity, and active erythropoiesis was also observed in these structures.
  
  
  CONCLUSIONS
  These results indicate that primitive erythropoiesis occurs in adult mice to rescue critical anemia because primitive erythropoiesis does not require macrophages as stroma whereas macrophages play a pivotal role in definitive erythropoiesis even outside the medulla. The cells expressing embryonic hemoglobin in this study were similar to primitive erythrocytes, indicating the possibility that yolk sac-derived primitive erythroid cells may persist into adulthood in mice.
authors.affiliation: Department of Oral Anatomy and Developmental Biology, School of Dentistry, Showa University, 1-5-8 Hatanodai, Shinagawa-ku, Tokyo, 142-8555 Japan.
authors.forename: Hirotada
authors.initials: H
authors.lastname: Otsuka
authors.affiliation: Department of Oral Anatomy and Developmental Biology, School of Dentistry, Showa University, 1-5-8 Hatanodai, Shinagawa-ku, Tokyo, 142-8555 Japan.
authors.forename: Jiro
authors.initials: J
authors.lastname: Takito
authors.affiliation: Division of Molecular Regulation, Graduate School of Dentistry, Tohoku University, 4-1 Seiryo-machi, Aoba-ku, Sendai, 980-8575 Japan.
authors.forename: Yasuo
authors.initials: Y
authors.lastname: Endo
authors.affiliation: Faculty of Pharmacy, International University of Health and Welfare, 2600-1 Kitakanamaru, Otawara-shi, Tochigi 324-8501 Japan.
authors.forename: Hideki
authors.initials: H
authors.lastname: Yagi
authors.affiliation: Department of Veterinary Anatomy, Nippon Veterinary and Animal Science University, 1-7-1 Kyonan-cho, Musashino-shi, Tokyo 180-8602 Japan.
authors.forename: Satoshi
authors.initials: S
authors.lastname: Soeta
authors.affiliation: Department of Oral Anatomy and Developmental Biology, School of Dentistry, Showa University, 1-5-8 Hatanodai, Shinagawa-ku, Tokyo, 142-8555 Japan.
authors.forename: Nobuaki
authors.initials: N
authors.lastname: Yanagisawa
authors.affiliation: Department of Oral Anatomy and Developmental Biology, School of Dentistry, Showa University, 1-5-8 Hatanodai, Shinagawa-ku, Tokyo, 142-8555 Japan.
authors.forename: Naoko
authors.initials: N
authors.lastname: Nonaka
authors.affiliation: Department of Oral Anatomy and Developmental Biology, School of Dentistry, Showa University, 1-5-8 Hatanodai, Shinagawa-ku, Tokyo, 142-8555 Japan.
authors.forename: Masanori
authors.initials: M
authors.lastname: Nakamura
keywords: Embryonic hemoglobin; Extramedullary erythropoiesis; KLF1; Nitrogen-containing bisphosphonate (NBP); Nucleated erythrocyte