Our principal hypotheses are that many as yet unidentified morphogens/growth factors control fetal and adult hematopoietic stem cell (HSC) survival and proliferation, and that genetic modifications of stromal cell lines can enhance their ability to support HSC expansion in culture without undergoing differentiation to lineage-restricted progenitors. We identified several potentially important secreted proteins specifically expressed in AFT024, a mouse fetal liver stromal cell line that maintains HSC stem cell activity: Pleiotrophin; Deltalike; the fibrillin-like protein T16; Cytokine Receptor-like Factor, and three members of the Proliferin gene family, Proliferin- 1; Mrp4, and proliferin- related protein (PRP). Using stable expression of siRNAs to block their production in AFT024 and AFT024 - FIt3L cells we will determine the function of these and other signaling proteins, such as M-CSF, in HSC expansion and differentiation. Using Fc fusion proteins we will determine whether HSCs have receptors for these novel growth/differentiation factors and whether their receptors might provide additional HSC- specific surface markers. Our preliminary data indicates positive effects of added Flt3L and IGF-2 on HSC maintenance and expansion in vitro. Thus, in parallel we will determine whether forced overexpression in AFT024 and AFT024 - Flt3L cells of several proteins including thrombopoietin; Stromal cell - derived factor 1; stem cell factor; IL-6; 3 Wnts; and IGF-2 enhances their ability to support expansion of HSCs in culture. As appropriate knock- out mice selectively missing one or more of these factors will be made and analyzed for HSCs and hematopoiesis. By comparing the ability of other stromal cell lines, including OP-9, MS-5, and S 17, to support maintenance and expansion of fetal liver and adult bone marrow HSCs, together with our existing transcriptional profiling data, we should identify other secreted/ surface proteins that potentially affect HSC expansion or differentiation positively or negatively, and in later years will test their role in HSC biology. Our long- term aim is to engineer a cloned line that supports robust, continuous expansion of fetal liver or bone marrow HSCs in culture. In continued collaboration with Prof. George Daley, we will test the ability of our novel growth factors/morphogens and genetically altered stromal cell lines to support the generation of transplantable HSCs from cultured human and mouse ES cell lines.

National Institute of Health (NIH)
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Research Project (R01)
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Hematopoiesis Study Section (HP)
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Bishop, Terry Rogers
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Whitehead Institute for Biomedical Research
United States
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Chou, Song; Flygare, Johan; Lodish, Harvey F (2013) Fetal hepatic progenitors support long-term expansion of hematopoietic stem cells. Exp Hematol 41:479-490.e4
Hattangadi, Shilpa M; Wong, Piu; Zhang, Lingbo et al. (2011) From stem cell to red cell: regulation of erythropoiesis at multiple levels by multiple proteins, RNAs, and chromatin modifications. Blood 118:6258-68
Bousquet, Marina; Lodish, Harvey F (2011) MicroRNAs: the primary cause or a determinant of progression in leukemia? Expert Rev Hematol 4:121-3
Khoury, Maroun; Drake, Adam; Chen, Qingfeng et al. (2011) Mesenchymal stem cells secreting angiopoietin-like-5 support efficient expansion of human hematopoietic stem cells without compromising their repopulating potential. Stem Cells Dev 20:1371-81
Chou, Song; Chu, Pat; Hwang, William et al. (2010) Expansion of human cord blood hematopoietic stem cells for transplantation. Cell Stem Cell 7:427-8
Chou, Song; Lodish, Harvey F (2010) Fetal liver hepatic progenitors are supportive stromal cells for hematopoietic stem cells. Proc Natl Acad Sci U S A 107:7799-804
Lodish, Harvey; Flygare, Johan; Chou, Song (2010) From stem cell to erythroblast: regulation of red cell production at multiple levels by multiple hormones. IUBMB Life 62:492-6
Zhang, Cheng Cheng; Kaba, Megan; Iizuka, Satoru et al. (2008) Angiopoietin-like 5 and IGFBP2 stimulate ex vivo expansion of human cord blood hematopoietic stem cells as assayed by NOD/SCID transplantation. Blood 111:3415-23
Huynh, Hoangdinh; Iizuka, Satoru; Kaba, Megan et al. (2008) Insulin-like growth factor-binding protein 2 secreted by a tumorigenic cell line supports ex vivo expansion of mouse hematopoietic stem cells. Stem Cells 26:1628-35
Zhang, Cheng C; Lodish, Harvey F (2008) Cytokines regulating hematopoietic stem cell function. Curr Opin Hematol 15:307-11

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