Adrenomedullin and Endothelin-1: Two New Growth Factors for In Vitro Expansion of Hematopoietic Cord Blood Cells

Abstract

Peptides derived from pro-adrenomedullin (AM) and endothelin (ET) establish two families of regulatory factors that are almost ubiquitously expressed in human tissues. Recently, some studies demonstrated that AM is expressed by all terminal cells of the lymphoid and myeloid pathways, while endothelin-1 (ET-1) is expressed and released by normal bone marrow platelet precursor cells. Hence, we investigated the inductive capacity of these two factors on clonal growth of hematopoietic cord blood stem cells (CBSCs), culturing whole cord blood, mononucleated cells and CD34⁺38⁻ subset population in the presence of high concentrations of AM, ET-1 (5 × 10⁻8 M) and their inhibitors (and 5 × 10⁻7 M). Moreover, we performed CBSCs middle-term expansion in a stroma-free liquid culture medium with AM, ET-1, and three different cytokine combinations. After 2 weeks of incubation, aliquots of expanded-cell suspensions were seeded on semisolid medium and clonogenic tests were carried out by counting the number of colony-forming units (CFUs) after 14 days of culture. Results obtained with these experiments demonstrated that: AM markedly enhanced CFUs growth in clonogenic tests carried out with whole cord blood, mononucleated cord blood cells, and with purified CD34⁺CD38⁻ cells population, whereas ET-1 increased only clonogenic growth of the last population subset; AM seemed to act via AM_22–52-sensitive receptors with an autocrine mechanism; both peptides we considered induced CBSCs expansion in middle-term expansion culture system, and they acted in a synergistic way with exogenous cytokines. These data suggest, for the first time, the role of AM on hematopoiesis regulation and the inductive effect of ET-1 on more uncommitted progenitor cells. We conclude that AM and ET-1 can be considered novel promising molecules to utilize, in association with cytokines, as CBSCs proexpansive factors in the clinical use of cord blood as an alternative source of stem cells for allogeneic transplantation.