Jiahui Zeng1, Danying Yi1, Wencui Sun1, Yuanlin Liu1, Jing Chang1, Lijiao Zhu1, Yonggang Zhang1, Xu Pan1, Yong Dong1, Ya Zhou1, Mowen Lai1, Guohui Bian1, Qiongxiu Zhou1, Jiaxin Liu1, Bo Chen2, Feng Ma3,4,5. 1. Chinese Academy of Medical Sciences & Peking Union Medical College (CAMS & PUMC), Institute of Blood Transfusion, No. 26, Huacai Road, Longtan Industry Park, Chenghua District, Chengdu, 610052, China. 2. Chinese Academy of Medical Sciences & Peking Union Medical College (CAMS & PUMC), Institute of Blood Transfusion, No. 26, Huacai Road, Longtan Industry Park, Chenghua District, Chengdu, 610052, China. 18980999910@163.com. 3. Chinese Academy of Medical Sciences & Peking Union Medical College (CAMS & PUMC), Institute of Blood Transfusion, No. 26, Huacai Road, Longtan Industry Park, Chenghua District, Chengdu, 610052, China. mafeng@hotmail.co.jp. 4. State Key Laboratory of Biotherapy, Sichuan University, Chengdu, 610065, China. mafeng@hotmail.co.jp. 5. State Key Laboratory of Experimental Hematology, CAMS & PUMC, Tianjin, 300020, China. mafeng@hotmail.co.jp.
Abstract
BACKGROUND: The HOX genes are master regulators of embryogenesis that are also involved in hematopoiesis. HOXA9 belongs to a cluster of HOX genes that play extensively studied roles in hematopoiesis and leukemogenesis. METHODS: We established HOXA9-inducible human embryonic stem cells (HOXA9/hESCs) with normal pluripotency and potential for hematopoiesis, which could be used to analyze gene function with high accuracy. HOXA9/hESCs co-cultured with aorta-gonad-mesonephros-derived stromal cells (AGM-S3) were induced to overexpress HOXA9 with doxycycline (DOX) at various times after hematopoiesis started and then subjected to flow cytometry. RESULTS: Induction of HOXA9 from Day 4 (D4) or later notably promoted hematopoiesis and also increased the production of CD34+ cells and derived populations. The potential for myelogenesis was significantly elevated while the potential for erythrogenesis was significantly reduced. At D14, a significant promotion of S phase was observed in green fluorescent protein positive (GFP+) cells overexpressing HOXA9. NF-κB signaling was also up-regulated at D14 following induction of HOXA9 on D4. All of these effects could be counteracted by addition of an NF-κB inhibitor or siRNA against NFKB1 along with DOX. CONCLUSIONS: Overexpression of HOXA9 starting at D4 or later during hematopoiesis significantly promoted hematopoiesis and the production of myeloid progenitors while reduced the production of erythroid progenitors, indicating that HOXA9 plays a key role in hematopoiesis and differentiation of hematopoietic lineages.
BACKGROUND: The HOX genes are master regulators of embryogenesis that are also involved in hematopoiesis. HOXA9 belongs to a cluster of HOX genes that play extensively studied roles in hematopoiesis and leukemogenesis. METHODS: We established HOXA9-inducible human embryonic stem cells (HOXA9/hESCs) with normal pluripotency and potential for hematopoiesis, which could be used to analyze gene function with high accuracy. HOXA9/hESCs co-cultured with aorta-gonad-mesonephros-derived stromal cells (AGM-S3) were induced to overexpress HOXA9 with doxycycline (DOX) at various times after hematopoiesis started and then subjected to flow cytometry. RESULTS: Induction of HOXA9 from Day 4 (D4) or later notably promoted hematopoiesis and also increased the production of CD34+ cells and derived populations. The potential for myelogenesis was significantly elevated while the potential for erythrogenesis was significantly reduced. At D14, a significant promotion of S phase was observed in green fluorescent protein positive (GFP+) cells overexpressing HOXA9. NF-κB signaling was also up-regulated at D14 following induction of HOXA9 on D4. All of these effects could be counteracted by addition of an NF-κB inhibitor or siRNA against NFKB1 along with DOX. CONCLUSIONS: Overexpression of HOXA9 starting at D4 or later during hematopoiesis significantly promoted hematopoiesis and the production of myeloid progenitors while reduced the production of erythroid progenitors, indicating that HOXA9 plays a key role in hematopoiesis and differentiation of hematopoietic lineages.
Entities:
Keywords:
Differentiation of hematopoietic lineages; HOXA9; Hematopoiesis; Human embryonic stem cells (hESCs); Inducible expression system; Mesoderm; Tet-on
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