Fig. 4. The effects of AVP or d(Leu4Lys8)VP, a specific AVPR1B agonist, on anemic rodents. The effects of AVP or d(Leu4Lys8)VP, a specific AVPR1B agonist,

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Fig. 4. The effects of AVP or d(Leu4Lys8)VP, a specific AVPR1B agonist, on anemic rodents. The effects of AVP or d(Leu4Lys8)VP, a specific AVPR1B agonist, on anemic rodents. (A) Avpr1a, Avpr1b, and Avpr2 receptor expression in fluorescence-activated cell–sorted mouse bone marrow (BM) lin−Sca-1+c-kit+ (LSK) cells. Lane 1, 100-bp marker; lane 2, RT-PCR product; lane 3, no template. (B) Flow cytometry sorting of mouse bone marrow erythroid lineage cells using CD44 and TER119 markers into four populations. Population I, proerythroblast; population II, basophilic; population III, polychromatic; population IV, orthochromatic. Numbers represent the percentage of total TER119-positive cells. n = 4 mice, C57BL/6. (C) Gene expression changes of Avpr1a, Avpr1b, and Avpr2 receptors in four different populations of bone marrow erythroid lineage cells. Error bars represent SEM; n = 4 mice, C57BL/6. TBP, TATA box–binding protein. (D) C57BL/6 mice were injected with AVP (100 μg/kg), and after 16 hours, the erythroid progenitors in the bone marrow and spleen were analyzed with flow cytometry. Data and error bars are means + SD; n = 5 mice per group; **P < 0.01. (E) C57BL/6 mice were hemorrhaged and injected once with AVP (100 μg/kg) or AVPR1B agonist (100 μg/kg). Hematocrit and corrected reticulocytes were measured on the days indicated. After hemorrhage, both AVP and d(Leu4Lys8)VP (AVPR1B agonist) significantly improved recovery times. Data and error bars are means + SD; n = 6 mice per group. *P < 0.05, **P < 0.01, ***P < 0.001. (F) C57BL/6 mice received whole-body irradiation (440 centigrays; two times within 12 hours) and were injected with AVP (100 μg/kg). Hematocrit and corrected reticulocytes were measured on the days indicated. After irradiation of the bone marrow, AVP administration resulted in a significantly higher hematocrit and reticulocyte values. Data and error bars are means + SD; n = 6 mice per group. *P < 0.05, ***P < 0.001. (G) C57BL/6 mice were splenectomized and allowed to recover for 1 month. The mice were subjected to hemorrhage and immediately injected with AVP (100 μg/kg) or AVPR1B agonist (100 μg/kg). Hematocrit and corrected reticulocytes were measured on the days indicated. In splenectomized mice, AVP still promoted recovery from anemia. Data and error bars are means + SD; n = 6 mice per group. **P < 0.01; ***P < 0.001. (H) Mice were sublethally irradiated and then given either wild-type (WT) or Avpr1b-deficient bone marrow. Two months later, they had 10 to 20% of their blood withdrawn or were treated with phenylhydrazine. Four days after hemorrhage, there was a significant increase in blood-corrected reticulocyte counts in animals given wild-type versus receptor-deficient bone marrow. This was not reflected in an increase in hematocrit. Six to 14 days after the phenylhydrazine treatment, corrected reticulocyte counts were significantly elevated in mice given wild-type bone marrow versus receptor-deficient bone marrow. On day 6, the hematocrit of the wild-type mice was also higher. Data and error bars are means + SD; n = 6 mice per group. *P < 0.05, **P < 0.01. (I) C57BL/6 mice were hemorrhaged (this sample was considered as the baseline) and injected with phosphate-buffered saline (PBS) or EPO (50 U/kg) or AVP (100 μg/kg). Hematocrits were measured at the time points indicated. AVP treatment significantly increased hematocrit by 12 hours compared to EPO or vehicle (PBS) treatment. Error bars represent SD; n = 6 mice per group. **P < 0.01, ***P < 0.001. (J) C57BL/6 mice were hemorrhaged and injected with either 100 μg of EPO-neutralizing antibody (Ab) or its isotype (ISO) control, with or without AVP (100 μg/kg). Hematocrits were measured at the time points indicated. Data and error bars are means + SD; n = 5 mice per group. *P < 0.05, ***P < 0.001. Balázs Mayer et al., Sci Transl Med 2017;9:eaao1632 Published by AAAS