Enhancing Erythropoiesis by a Phytoestrogen Diarylheptanoid from Curcuma comosa.

Erythropoietin (Epo) is widely used for the treatment of anemia; however, non-hematopoietic effects and cancer risk limit its clinical applications. Therefore, alternative molecules to improve erythropoiesis in anemia patients are urgently needed. Here, we investigated the potential effects of a phytoestrogen diarylheptanoid (3R)-1,7-diphenyl-(4E,6E)-4,6-heptadien-3-ol, (ASPP 049) isolated from Curcuma comosa on promoting erythropoiesis. Treatment with C. comosa extract improved anemia symptoms demonstrated by increasing red blood cell numbers, hematocrit, and hemoglobin content in anemic mice. In addition, ASPP 049, the major compound isolated from C. comosa, enhanced the suboptimal Epo dosages to improve erythroid cell differentiation from hematopoietic stem cells, which was inhibited by the estrogen receptor (ER) antagonist, ICI 182,780. Moreover, the ASPP 049-activated Epo-Epo receptor (EpoR) complex subsequently induced phosphorylation of EpoR-mediated erythropoiesis pathways: STAT5, MAPK/ERK, and PI3K/AKT in Epo-sensitive UT-7 cells. Taken together, these results suggest that C. comosa extract and ASPP 049 increased erythropoiesis through ER- and EpoR-mediated signaling cascades. Our findings provide insight into the specific interaction between a phytoestrogen diarylheptanoid and Epo-EpoR in a hematopoietic system for the potential treatment of anemia.

Iron-loaded transferrin potentiates erythropoietin effects on erythroblast proliferation and survival: a novel role through transferrin receptors.

Red blood cell production, or erythropoiesis, is a proliferative process that requires tight regulation. Erythropoietin (Epo) is a glycoprotein cytokine that plays a major role in erythropoiesis by triggering erythroid progenitors/precursors of varying sensitivity. The concentration of Epo in bone marrow is hypothesized to be suboptimal, and the survival of erythroid cells has been suggested to depend on Epo sensitivity. However, the key factors that control Epo sensitivity remain unknown. Two types of transferrin receptors (TfRs), TfR1 and TfR2, are known to play a role in iron uptake in erythroid cells. Here, we hypothesized that TfRs may additionally modulate Epo sensitivity during erythropoiesis by modulating Epo receptor (EpoR) signaling. Using an Epo-sensitive UT-7 (UT7/Epo) erythroid cell and human erythroid progenitor cell models, we report that iron-loaded transferrin, that is, holo-transferrin (holo-Tf), synergizes with suboptimal Epo levels to improve erythroid cell survival, proliferation, and differentiation. This is accomplished via the major signaling pathways of erythropoiesis, which include signal transducer and activator of transcription 5 (STAT5), mitogen-activated protein kinase/extracellular signal-regulated kinase (MAPK/ERK), and phosphoinositide-3-kinase (PI3K)/AKT. Furthermore, we found that this cooperation is improved by, but does not require, the internalization of TfR1. Interestingly, we observed that loss of TfR2 stabilizes EpoR levels and abolishes the beneficial effects of holo-Tf. Overall, these data reveal novel signaling properties of TfRs, which involve the regulation of erythropoiesis through EpoR signaling.

Ex vivo expansion and functional activity preservation of adult hematopoietic stem cells by a diarylheptanoid from Curcuma comosa.

Hematopoietic stem cells (HSCs, CD34+ cells) have shown therapeutic efficacy for transplantation in various hematological disorders. However, a large quantity of HSCs is required for transplantation. Therefore, strategies to increase HSC numbers and preserve HSC functions through ex vivo culture are critically required. Here, we report that expansion medium supplemented with ASPP 049, a diarylheptanoid isolated from Curcuma comosa, and a cocktail of cytokines markedly increased numbers of adult CD34+ cells. Interestingly, phenotypically defined primitive HSCs (CD34+CD38-CD90+) were significantly increased under ASPP 049 treatment relative to control. ASPP 049 treatment also improved two functional properties of HSCs, as evidenced by an increased number of CD34+CD38- cells in secondary culture (self-renewal) and the growth of colony-forming units as assessed by colony formation assay (multilineage differentiation). Transplantation of cultured CD34+ cells into immunodeficient mice demonstrated the long-term reconstitution and differentiation ability of ASPP 049-expanded cells. RNA sequencing and KEGG analysis revealed that Hippo signaling was the most likely pathway involved in the effects of ASPP 049. These results suggest that ASPP 049 improved ex vivo expansion and functional preservation of expanded HSCs. Our findings provide a rationale for the use of ASPP 049 to grow HSCs prior to hematological disease treatment.