Abstract
Ecotropic viral integration site-1 (EVI1) has a critical role in normal and malignant hematopoiesis. Since we previously identified high expression of calcitonin receptor like receptor (CRLR) in acute myeloid leukemia (AML) with high EVI1 expression, we here characterized the function of CRLR in hematopoiesis. Since higher expression of CRLR and receptor activity modifying protein 1 (RAMP1) was identified in immature hematopoietic bone marrow (BM) cells, we focused on calcitonin gene-related peptide (CGRP), a specific ligand for the CRLR/RAMP1 complex. To elucidate the role of CGRP in hematopoiesis, Ramp1-deficient (Ramp1−/−) mice were used. The steady-state hematopoiesis was almost maintained in Ramp1−/− mice; however, the BM repopulation capacity of Ramp1−/− mice was significantly decreased, and the transplanted Ramp1−/− BM mononuclear cells had low proliferation capacity with enhanced reactive oxygen species (ROS) production and cell apoptosis. Thus, CGRP is important for maintaining hematopoiesis during temporal exposures with proliferative stress. Moreover, continuous CGRP exposure to mice for two weeks induced a reduction in the number of BM immature hematopoietic cells along with differentiated myeloid cells. Since CGRP is known to be increased under inflammatory conditions to regulate immune responses, hematopoietic exhaustion by continuous CGRP secretion under chronic inflammatory conditions is probably one of the important mechanisms of anti-inflammatory responses.
Highlights
Hematopoietic stem and progenitor cells (HSPCs) stay in the bone marrow (BM) niches that controls survival, self-renewal, or differentiation in the BM1–8
calcitonin generelated peptide (CGRP) is a ligand of Crlr/Ramp[1] in murine hematopoietic cells and the hematopoiesis of Ramp1deficient mice is maintained under steady-state condition
To determine whether Crlr is expressed in normal murine bone marrow mononuclear cells (BMMNCs), several BM cell fractions [lineage marker (Lin)− Sca-1+ c-kit+ (LSK), common myeloid progenitors (CMP) (Lin− Sca-1− c-kit+ CD16/32− CD34+), granulocyte/monocyte progenitor (GMP) (Lin− Sca-1− c-kit+ CD16/32+ CD34+), megakaryocyte/erythroid progenitor (MEP) (Lin− Sca-1− c-kit+ CD16/32+ CD34+), myeloid (Gr-1+CD11b+), and lymphoid (B220+ for B lineage and CD3e+ for T lineage)] were separated from mouse BM by cell sorting using specific antibodies (Methods), and levels of Crlr mRNA and protein expression were determined in the BM fractions by real-time PCR (Fig. 1a,b) and flow cytometry (Fig. 1c), respectively
Summary
Hematopoietic stem and progenitor cells (HSPCs) stay in the BM niches that controls survival, self-renewal, or differentiation in the BM1–8. EVI1 plays an important role in maintaining hematopoietic stem cells (HSCs) and LSCs, the exact molecular functions of EVI1 in stem cells has not been elucidated[19,20,21]. We identified that RAMP1 is expressed in hematopoietic progenitor cells, which led us to speculate that CGRP might be a specific ligand for CRLR in hematopoietic cells. Here, we investigated how CGRP-mediated activation of the CRLR/RAMP1 signaling pathway affects hematopoiesis using Ramp1−/− mice. The hematopoiesis of Ramp1−/− mice under steady-state conditions was not affected, which was revealed by examining their hematopoietic progenitor cells in the BM and differentiated blood cells in the peripheral blood (PB). The CGRP-CRLR/RAMP1 signaling pathway might play an important role in hematopoiesis under proliferative stress conditions. Given that long-term CGRP administration to mice decreased the numbers of most of immature hematopoietic cells and differentiated blood cells, it is suggested that continuous CGRP secretion under inflammation and chronic disease may be involved in the hematopoietic exhaustion
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